Coproantigen Test Research Articles

Investigation of Factors Associated with Subclinical Infections of Giardia duodenalis and Cryptosporidium canis in Kennel-Housed Dogs (Canis lupus familiaris).

Giardia duodenalis and Cryptosporidium spp. are zoonotic protozoal pathogens, spread by a fecal-oral route, which can infect a wide range of hosts including but not limited to dogs and humans. Giardia was recently estimated to be present in 37% to 50% of kennel-housed dogs. Cryptosporidium infections in kennel-housed dogs have been reported in 7% to 21% of the population. The goal of this study was to define demographic factors and fecal scores associated with positive screening test cases of Giardia and Cryptosporidium in kennel-housed laboratory dogs in the state of Texas. Fecal samples were collected from 153 clinically normal laboratory dogs at an academic research facility and a local laboratory dog supplier. We used 3 diagnostic tests evaluated in parallel to determine test positivity to each organism: a human point-of-care coproantigen test, a direct immunofluorescent assay, and an in-house polymerase chain reaction. Dogs were significantly more likely to test positive for Giardia (45%) than Cryptosporidium (7%) (P < 0.01). Dogs that were 18 mo of age or younger had 3 times the odds (P = 0.009) of subclinical Giardia infection compared with older dogs. We found no significant relationship between age and Cryptosporidium prevalence. Dogs with hard feces (fecal score 1-2) at the time of screening had 0.34 times lower odds ( P = 0.049) of testing positive for Giardia than dogs with normal feces, but no statistically significant relationship was found between fecal score and Cryptosporidium -positive test status. With these findings, we demonstrated the value of considering age and fecal score when choosing which dogs to screen for subclinical Giardia. Additional studies with larger sample sizes should be conducted to determine the relationship between age and fecal score and subclinical Cryptosporidium infection.

Efficacy of flukicides against Fasciola hepatica and first report of triclabendazole resistance on German sheep farms

Fasciola hepatica infections lead to severe health problems and production losses in sheep farming, if not treated effectively. Triclabendazole has been used extensively over decades due to its unique efficacy range against all definitive hostfluke stages but published data about the susceptibility of F. hepatica to anthelmintics in Germany are lacking. This study aimed to identify current F. hepatica infections in German sheep flocks by coproscopic examinations and to evaluate the efficacy of anthelmintics with a focus on triclabendazole in a field study conducted from 2020 to 2022. Initial screening included 71 sheep farms, many of them with known history of fasciolosis. In this highly biased sample set, the frequency of F. hepatica infection at individual sheep and farm level were 12.8% and 35.2%, respectively. Additionally, eggs of Paramphistominae were found at frequencies of 4.8% and 15.5% at individual sheep and farm level, respectively. Due to low egg shedding intensity, faecal egg count reduction (FECR) tests could only be conducted on a few farms. The efficacy of triclabendazole was tested on 11 farms and albendazole on one farm, including 3–53 sheep/farm. Individual faecal samples were collected before and two weeks after treatment to evaluate the FECR using the sedimentation or FLUKEFINDER® or a modified FLUKEFINDER® method. On all farms a coproantigen reduction test was conducted in parallel. Lacking efficacy of triclabendazole even at double dosage was shown on one farm associated with a high number of animal losses due to acute fasciolosis. On this farm, the Fasciola miracidium development test was additionally performed, revealing a high in vitro ovicidal activity of albendazole while closantel was effective in vivo. On all other farms, sufficient efficacy of triclabendazole was observed. In conclusion, triclabendazole resistance appears not to be widespread on German sheep farms but, when present, can have serious effects on animal health.

Diagnosis of canine intestinal parasites: Improved detection of Dipylidium caninum infection through coproantigen testing

Intestinal parasites, including cestodes like Dipylidium caninum, are common in dogs in the United States of America (USA), but fecal flotation consistently, and, at times, dramatically, fails to identify many of these infections. To determine the extent to which including coproantigen testing for D. caninum would improve the identification of dogs infected with this cestode, we evaluated fecal samples from 877 dogs (589 pet and 288 from municipal shelters) from six USA states using zinc sulfate (specific gravity 1.24) fecal flotation with centrifugation along with coproantigen detection for Giardia sp., hookworms, ascarids, and Trichuris vulpis. For D. caninum, PCR of perianal swabs was included. Intestinal parasite infections were identified, using centrifugal fecal flotation or coproantigen, in 265 dogs (13.2 % pet, 64.9 % shelter). Dipylidium caninum infection was detected in 5.6 % of dogs with the combination of coproantigen and centrifugal fecal flotation, and 7.3 % of dogs when perianal swab results were included; prevalence varied by diagnostic method, population, and geographic region. In pet dogs, D. caninum infection was identified by fecal flotation (0), coproantigen (2.2 %), or perianal swabs (1.2 %). The same methods revealed infection in 0.3 %, 12.5 %, and 11.1 % of shelter dogs, respectively. Frequent use of praziquantel in shelter dogs (116/288; 40.3 %) may have reduced prevalence. Positive and negative agreement of D. caninum coproantigen with perianal swab PCR in pet dogs was 85.7 % and 98.8 %, respectively. Multiple logistic regression analysis accounting for region, population, and age found D. caninum infection to be more common in shelter dogs relative to pet (adjusted OR 4.91 [2.48, 10.24]) and in the Southcentral and Southeast regions relative to North (adjusted OR 9.59 [1.92, 174.13] and 17.69 [3.67, 318.09] respectively). Coproantigen testing also enhanced the detection of other intestinal parasites over fecal flotation alone, including Giardia sp. (14.7 % vs 3.3 %), hookworms (13.8 % vs 8.4 %), ascarids (2.9 % vs 2.2 %), and T. vulpis (2.9 % vs 1.4 %). Together, these data indicate that the coproantigen assay employed increases detection of D. caninum infections several fold, supporting the use of this test in clinical practice, and add to a growing body of research documenting enhanced diagnosis through implementation of multiple laboratory-based methods.

Diagnostic Value of Coproantigen for Detection of Giardia Infection in Stunted Children

Background: Giardiasis is a protozoa infection caused by Giardia intestinalis, which commonly infects children, impairing children’s growth, development, and cognitive function. Standard diagnosis is carried out by microscopic examination of stool. This study aimed to evaluate coproantigen examination in stunted children compared to microscopic examination.Methods: A cross-sectional study was performed on stools collected from a survey among stunted children in Bandung in 2019. Stools were preserved in 10% formaldehyde and kept at -20oC until used. Direct microscopy examination with 2% lugol solution and coproantigen ELISA test using Giardia Cryptosporidium (combo test) coproantigen test kit were performed in Parasitology Laboratory Faculty of Medicine, Universitas Indonesia.Result: A total of 99 stools originated from stunted children aged 2-6 years. with boys predominant (52.5%). Microscopic examination showed that 12.1% (12/99) of the children were harboring intestinal parasites, such as the protozoa Giardia intestinalis, Blastocystis spp, and Entamoeba coli (E. coli). Giardia was the primary infection (9.1%), of which single Giardia infection (n = 8) and mixed infection of Giardia and Blastocystis spp (n = 1). Interestingly, coproantigen examination resulted in 6 positive samples, and 4 samples agreed with the microscopy result. With a sensitivity of 44.4% and a specificity of 97.7%. The positive and negative predictive values were 66.7% and 94.7%, respectively.Conclusion: A moderate prevalence of Giardia in stunted children in Bandung regency has been observed. The combo coproantigen test method has high specificity and is suitable for use as a confirmation test to exclude Giardia infection.

Comparison of Kato-Katz, PCR and coproantigen for the diagnosis of Taenia solium taeniasis.

Four methods were compared for the diagnosis of human taeniasis caused by Taenia solium. Fecal samples from persons living in a T. solium endemic region of Madagascar were examined for taeniid eggs by the Kato–Katz method. Subsequently, samples positive (n = 16) and negative (n = 200) for T. solium eggs were examined by (i) amplification of the fragment of small subunit of the mitochondrial ribosomal RNA (rrnS) gene using conventional polymerase chain reaction (PCR) and (ii) a nested PCR of a fragment of the T. solium Tso31 gene. Additionally, 12 egg-positive and all egg-negative samples were tested for coproantigen detection. A further 9 egg-positive fecal samples were examined using both PCRs. Of the 12 egg-positive samples tested by PCRs and coproantigen methods, 9 (75%) were positive by rrnS PCR, 3 (25%) using Tso31-nested PCR and 9 (75%) by coproantigen testing. None of the 200 egg-negative fecal samples was positive in either rrnS or Tso31-nested PCR. Twenty of the 25 egg-positive samples (80%) were positive in rrnS PCR, and DNA sequencing of PCR amplicons was obtained from 18 samples, all confirmed to be T. solium. Twelve of the 25 egg-positive samples (48%) were positive in the Tso31-nested PCR, all of which were also positive by rrnS PCR. It is suggested that species-specific diagnosis of T. solium taeniasis may be achieved by either coprological examination to detect eggs or coproantigen testing, followed by rrnS PCR and DNA sequencing to confirm the tapeworm species in egg-positive or coproantigen-positive samples.

Immunoassay for detection of Dipylidium caninum coproantigen in dogs and cats.

Dipylidium caninum infections in dogs and cats are underestimated because of a lack of proglottid observations and poor recovery of parasite elements by centrifugal flotation. We developed an immunoassay that employs a pair of monoclonal antibodies to capture D. caninum-specific coproantigen in fecal extracts from dogs and cats. Real-time PCR for D. caninum DNA in perianal swabs and observation of proglottids were used as reference methods. In 6 experimentally infected dogs, parasite DNA, coproantigen, and proglottid segments were first detected at 22, 23, and 26 d post-infection, respectively. Praziquantel treatment of 3 experimentally infected dogs resulted in the elimination of both coproantigen and proglottid shedding within 1-5 d post-treatment; however, parasite DNA persisted for 14 d. Immunohistochemistry on immature and mature tapeworm segments using an antibody against the coproantigen supports the premise that the antigen is produced in mature segments. We assessed the performance of our coproantigen test in natural infections in 78 dogs from a flea-endemic area. Of the 12 antigen-positive samples, 11 were confirmed with a positive PCR test and/or proglottid observation. Finally, we evaluated a convenience sample set of 730 canine and 163 feline fecal samples obtained from a commercial diagnostic laboratory; D. caninum antigen was detected in 4.1% of the canine and 12.9% of the feline samples, whereas parasite elements were observed in only 0.028% of samples. Our coproantigen immunoassay provides a sensitive method for the detection of D. caninum infection in dogs and cats.

A 'One Health' perspective of Africa-wide distribution and prevalence of Giardia species in humans, animals and waterbodies: a systematic review and meta-analysis.

Giardiasis, caused by Giardia duodenalis, is a leading cause of diarrhoea in resource-poor countries. To gain a better insight into the epidemiology of Giardia in Africa, we undertook a robust study to comprehend the distribution and prevalence of Giardia infection in humans, animals and their dispersal in the environment. Our protocol was registered with PROSPERO (registration number CRD42022317653). Deep literature search from 5 electronic databases, namely, AJOL, Google scholar, PubMed, ScienceDirect and Springer Link was performed using relevant keywords. Meta-analysis was performed using a random-effects model and heterogeneity among studies was evaluated using Cochran's Q and the I2-statistic. More than 500 eligible studies published from 1 January 1980 until 22 March 2022 were retrieved. In humans, exactly 48 124 Giardia spp. infection cases were registered from the 494 014 stool samples examined resulting in a pooled prevalence estimate (PPE) of 8.8% using microscopy. Whereas copro-antigen tests and molecular diagnostic methods generated PPE of 14.3 and 19.5%, respectively, with HIV+ subjects and those with diarrhoeatic stool having infection rates of 5.0 and 12.3%, respectively. The PPE of Giardia spp. infection in animals using molecular methods was 15.6%, which was most prevalent in pigs (25.2%) with Nigeria registering the highest prevalence at 20.1%. The PPE of Giardia spp. contamination from waterbodies was 11.9% from a total of 7950 samples which were detected using microscopy, with Tunisia documenting the highest infection rate of 37.3%. This meta-analysis highlights the necessity of ‘One Health’ approach for consolidated epidemiological studies and control of giardiasis in the African continent.

Retrospective Survey of Dog and Cat Endoparasites in Ireland: Antigen Detection.

Endoparasites of dogs and cats, play an important role in both veterinary medicine and public health. Untreated and stray dogs and cats, in particular, play an important role in contaminating the environment with important zoonotic parasites. Thus, the aim of this study was to estimate the prevalence of intestinal parasites in stray dogs and cats using highly sensitive and specific copro-antigen tests. Archive faecal samples from previous surveys conducted between 2016−2019 from dogs (n = 789) and cats (n = 241) were included in this study. The IDEXX Fecal Dx™ antigen panel was used for the detection of Toxocara, hookworms, Trichuris and the SNAP™ Giardia antigen assay was used for the detection of Giardia infection. Giardia duodenalis was the most common parasite (26%, n = 205) detected in the dogs, followed by ascarids (17.6%, n = 139) and hookworms (5.3%, n = 42). Trichuris vulpis was only detected in 1 dog. Ascarids (23.2%, n = 56) was the most common parasite detected in the cats, followed by Giardia (12.9%, n = 31) and hookworms (n = 7, 2.9%). No whipworms were detected in cats. Overall, there was little difference in the positivity between sexes in both dogs and cats. However, in terms of age, adolescent dogs (<3 years) and kittens (<1 year) had the highest parasite prevalence overall, with G. duodenalis and ascarids being the most prevalent. This study shows a high prevalence of parasite infection in untreated and stray dogs and cats in the greater Dublin area in Ireland. Since they live in synanthropic conditions and can roam over vast distances they can contaminate public areas and pose a risk to both humans and owned pets that utilise these spaces. It is therefore important to raise public awareness and increase the knowledge on zoonotic parasites.

Evaluation of a commercial coproantigen immunoassay for the detection of Toxocara cati and Ancylostoma tubaeforme in cats and Uncinaria stenocephala in dogs

Coproantigen immunoassays (IDEXX Fecal Dx® antigen tests) were evaluated for their ability to identify Toxocara cati and Ancylostoma tubaeforme infections in cats and Uncinaria stenocephala infection in dogs. Five cats were experimentally infected with 500 embryonated eggs of T. cati, eight cats with 500 third-stage larvae (L3) of A. tubaeforme and seven dogs with 500 L3 of U. stenocephala. In addition to the three coproantigen tests, the course of infection was monitored by a combined sedimentation-flotation method with ZnSO4 as flotation medium (specific gravity: 1.28–1.30) and a modified McMaster method in case of copromicroscopically positive samples. Eggs of T. cati were first observed between 28 and 54 days post infection (dpi) in four of the five infected cats. In these four cats, positive roundworm coproantigen signals were obtained between 16 and 44 dpi. Positive coproantigen signal always preceded egg observations, but the interval varied between 6 and 30 days. Hookworm-specific positive coproantigen signals were detected in seven of the eight A. tubaeforme infected cats between 10 and 52 dpi, while consecutive egg excretion was observed in three cats between day 26 and 54 pi. Of these three, coproantigen signal preceded egg observation by 12 to 24 days. Four cats had positive coproantigen results in the absence of egg excretion, and one cat never achieved a positive result for egg or coproantigen. In six of seven U. stenocephala infected dogs, infection was confirmed by copromicroscopy between 16 and 24 dpi as well as for hookworm coproantigen between 10 and 14 dpi. Coproantigen signal was detected prior to egg observation by 2 to 14 days. No cross-reactions between the roundworm, hookworm und whipworm tests occurred in study animals. The results of this study demonstrate the ability of the coproantigen tests to detect the common roundworm and hookworm infections in cats and U. stenocephala infections in dogs as well as the ability to detect the prepatent stage of infection.

Canine nematode and Giardia spp. infections in dogs in Edmonton, Alberta, the “CANIDA” study

BackgroundCanine intestinal parasite prevalence may be influenced by geographical region, age, and health status of the dog. Behaviors such as predation, scavenging, or roaming as well as routine administration of anthelmintics also play a role. The purpose of this study was to evaluate fecal test results using zinc sulfate flotation by centrifugation combined with coproantigen testing directed at protein antigens excreted or secreted by hookworms (Ancylostoma spp. Uncinaria stenocephala), ascarids (Toxocara canis, Toxascaris spp. Baylisascaris spp.), whipworms (Trichuris vulpis), and Giardia spp. during active infection in owned dogs visiting dog parks in Western Canada.MethodsA total of 774 participants were recruited from Edmonton, Alberta, Canada. Canine fecal samples were collected from seven dedicated off-leash dog parks. Participating dog owners responded to a questionnaire regarding their dogs’ signalment, previous veterinary history, and use of parasite-preventive products. Fecal samples were tested using zinc sulfate centrifugation combined with coproantigen testing.ResultsThe overall prevalence of canine intestinal parasites in client-owned dogs was similar to previous studies conducted in the US. Mean age of dogs tested was 4 years, with puppies and older dogs having higher rates of infection than the mean. Fecal flotation centrifugation found 3.2% hookworm, ascarid, whipworm, and Giardia spp.-positive infections. Coproantigen testing identified 5.8% positive infections, including all of the above that were detected using fecal flotation centrifugation.ConclusionsCoproantigen testing detected more hookworm, ascarid, whipworm, and Giardia spp.-positive samples in addition to detecting all positive results found using fecal flotation centrifugation. Fecal flotation centrifugation combined with coproantigen testing improves sensitivity over flotation alone and may detect pre-patent or sub-clinical infections in dogs visiting public dog parks.Graphical

Application of Urine and Copro Antigen Assays after Primary Infection and Drug Treatment in an Experimental Opisthorchiasis Animal Model.

Infection by Opisthorchis viverrini causes significant health problems, including cholangiocarcinoma (CCA); thus control and elimination of this trematode is an important strategy for the reduction of CCA. Currently, urine and copro antigen detection is more sensitive than parasitological examination of the feces for the diagnosis of opisthorchiasis. Given limitations in human studies, we used an animal model to quantify the parasite antigen profiles in urine and feces in O. viverrine-infected hamsters, and postdrug treatment. The positive detections of O. viverrini antigen began from week 1 in urine and week 2 in feces after infection until week 28 of the study. The recoveries of O. viverrini worms were detected starting from week 1 and eggs of O. viverrini were detected in feces from week 3 after infection and remained detectable throughout the study period. There was a significant positive correlation of urine and copro antigen levels with the number of fecal egg counts (P < 0.01) and worm recovery (P < 0.01). In the drug-treatment experiment, treatment of infected hamsters with praziquantel significantly reduced worm burden, fecal egg output, and antigen in urine and feces compared with the untreated controls (P < 0.001). At 4 weeks posttreatment, the egg and worm reduction rates were 100% and 95.5%, respectively. The positive antigen detections in urine and feces corresponded with partial worm clearance from praziquantel treatment. This study demonstrated a direct link of urine and copro antigen tests with worms infecting the liver thereby reaffirming the reliability of urine and copro antigen assay in opisthorchiasis diagnosis.

A Comparative Analysis of Microscopy, Coproantigen Serology, and Nested Multiplex PCR in the Laboratory Diagnosis of Entamoeba histolytica Infection.

Objectives Amoebiasis is caused by the most common intestinal protozoan parasite Entamoeba histolytica . This parasite causes amoebic colitis, which is manifested by diarrhea, followed by dysentery. The laboratory diagnosis of intestinal amoebiasis in most cases is by microscopic examination of stool samples. Other nonroutine methods include coproantigen enzyme-linked immunosorbent assay (ELISA) from stool samples, serum ELISA for antibodies, stool culture, isoenzyme analysis, and polymerase chain reaction (PCR). The present study aimed to comparatively analyze the different diagnostic modalities used for the detection of E. histolytica from the stool sample of patients with intestinal amoebiasis. Materials and Methods This study was undertaken with 631 patients, during a period of 3 years, from January 2017 to December 2019. Stool specimen obtained from each patient was subjected to direct microscopic wet mount examination, coproantigen ELISA, and nested multiplex PCR, respectively. Results Out of all the patients tested, 5.2% were positive for E. histolytica. Among the positive cases, stool microscopy was positive in 3.17%, coproantigen ELISA was positive in 29 (4.6%) cases, and PCR was positive in 30 (4.75%) cases. Statistical Analysis The prevalence of E. histolytica infection was summarized as percentages. The three diagnostic tests done were statistically analyzed, taking microscopy as the gold standard. The agreement between techniques (microscopy, coproantigen ELISA, and PCR) was analyzed with kappa statistics. Sensitivity, specificity, positive predictive value, negative predictive value, and diagnostic accuracy were summarized as percentage with 95% confidence interval. Conclusion In all suspected amoebiasis cases, a combination of stool microscopy, coproantigen testing with molecular detection of the parasite offers the best approach to diagnosis of this parasitic infection.

Diagnosis of sheep fasciolosis caused by Fasciola hepatica using cathepsin L enzyme-linked immunosorbent assays (ELISA)

Fasciolosis, a global parasitic disease of agricultural livestock, is caused by the liver fluke Fasciola hepatica. Management and strategic control of fasciolosis on farms depends on early assessment of the extent of disease so that control measures can be implemented quickly. Traditionally, this has relied on the detection of eggs in the faeces of animals, a laborious method that lacks sensitivity, especially for sub-clinical infections, and identifies chronic infections only. Enzyme linked immunosorbent assays (ELISA) offer a quicker and more sensitive serological means of diagnosis that could detect early acute infection before significant liver damage occurs. The performance of three functionally-active recombinant forms of the major F. hepatica secreted cathepsins L, rFhCL1, rFhCL2, rFhCL3, and a cathepsin B, rFhCB3, were evaluated as antigens in an indirect ELISA to serologically diagnose liver fluke infection in experimentally and naturally infected sheep. rFhCL1 and rFhCL3 were the most effective of the four antigens detecting fasciolosis in sheep as early as three weeks after experimental infection, at least five weeks earlier than both coproantigen and faecal egg tests. In addition, the rFhCL1 and rFhCL3 ELISAs had a very low detection limit for liver fluke in lambs exposed to natural infection on pastures and thus could play a major role in the surveillance of farms and a ‘test and treat’ approach to disease management. Finally, antibodies to all three cathepsin L proteases remain high throughout chronic infection but decline rapidly after drug treatment with the flukicide, triclabendazole, implying that the test may be adapted to trace the effectiveness of drug treatment.

Prevalence of endoparasitic and viral infections in client-owned cats in metropolitan Bangkok, Thailand, and the risk factors associated with feline hookworm infections.

The prevalence of enteric parasites in cats in metropolitan Bangkok has not been updated in over 13years. The main objectives of this study include updating the prevalence of endoparasitism in client-owned cats, status of retroviral infections and determining the association between feline hookworm infection and possible risk factors. A total of 509 fecal samples were collected from client-owned cats in 2014-2015 and examined by a wet fecal mount technique. If additional sample remained, a PBS-ethyl acetate sedimentation was done (n=229), and ZnSO4 centrifugal flotation was also performed if there was sufficient remaining sample (n=105). At least one parasite was observed in 32.0% (163/509) of cats, with Ancylostoma being the most common intestinal parasite detected in 21.6% (110/509) of cats. Other parasitic infections detected by fecal examinations included Toxocara (6.9%; 35/509), Platynosomum (3.7%; 19/509), Cystoisospora (3.5%; 18/509), Taenia (2.9%; 15/509), Spirometra (1.6%; 8/509), Dipylidium (0.4%; 2/509), and Opisthorchis-like trematode (0.2%; 1/509). Examination for Giardia infection was conducted with the SNAP® Giardia Test, a coproantigen test, on a subset of the fecal samples (233/509) and revealed a positive result on 3.9% (9/233) of samples. Plasma samples were analyzed using the SNAP® Triple Test detecting antigens of Feline Leukemia Virus (FeLV) and Dirofilaria immitis while also detecting antibodies to Feline Immunodeficiency Virus (FIV). Antigens of FeLV and antibodies to FIV were found in 7.1% (19/269) and 5.2% (14/269) of cats, respectively. None of the cats were found to have circulating antigen of Dirofilaria immitis using this test. No association between retroviral and endoparasitic infections was found. From multivariable logistic regression examining associated factors, the ability of cats to access the outdoors (adjusted OR=3.22, 95% CI; 1.42-7.87) and having tapeworm segments or adult helminths in feces (adjusted OR=3.31, 95% CI; 1.34-8.21) were significantly associated with the finding of hookworm eggs in feces. This work presents the most up-to-date data on enteric feline parasite prevalence in the metropolitan Bangkok area from which fecal samples were directly collected from cats. Consequently, this study emphasizes that diagnosis of parasitic infections and the routine use of antiparasitic medications should be encouraged by veterinarians and to owners in order to reduce the reservoir of potentially zoonotic parasites.

Assessment of a Multiplex PCR for the Simultaneous Diagnosis of Intestinal Cryptosporidiosis and Microsporidiosis: Epidemiologic Report from a French Prospective Study

Microsporidiosis and cryptosporidiosis are associated with chronic diarrhea in immunocompromised patients. The objectives of this study were to: i) assess a multiplex quantitative PCR assay targeting Cryptosporidium spp and the microsporidian Enterocytozoon bieneusi and Encephalitozoon spp, and ii) provide an update on the epidemiology of these pathogens. A prospective study was conducted from January 2017 to January 2019. Performance of the assay was assessed, and all cryptosporidia and microsporidia isolates were genotyped. The sensitivity of the multiplex PCR method reached 1 copy/μL for each targeted pathogen. The sensitivity of co-proantigen testing in the diagnosis of cryptosporidiosis was 73%. The sensitivity of microscopy in the diagnosis of cryptosporidiosis was 64%, and microsporidiosis, 50%. Among the 456 patients included, 14 were positive for Cryptosporidium spp (4 different species); 5, for E. bieneusi; and 2, for Encephalitozoon intestinalis. The overall prevalence of cryptosporidia was 3.1%, and of microsporidia, 1.5%; in kidney transplant recipients (n= 82), corresponding values were 7.3% and 2.4% (6 and 2 patients), respectively. Two cases of E. intestinalis infection were diagnosed in children who had traveled to the tropics. This study is the first to assess a multiplex quantitative PCR method for the simultaneous diagnosis of intestinal microsporidiosis and cryptosporidiosis. The highest prevalences of both pathogens were observed in kidney transplant recipients.

Retrospective analysis of feline intestinal parasites: trends in testing positivity by age, USA geographical region and reason for veterinary visit

BackgroundThe goals of this retrospective study were to estimate parasite positivity in samples from cats using zinc sulfate fecal flotation by centrifugation (“centrifugation”) and coproantigen and examine trends with age, geographical region and reason for visit to veterinarian. Common methods of parasite detection, such as centrifugal flotation, passive flotation, or direct smear, may underrepresent the true prevalence of intestinal parasites in cats. Coproantigen testing detects more positive samples than traditional methods alone.MethodsFeline fecal test results from the continental USA containing results for fecal exams performed using centrifugation paired with coproantigen results for ascarid, hookworm, whipworm and Giardia were obtained from the database of a national commercial reference laboratory comprised of multiple regional sites.ResultsParasite positivity was highest in samples from young cats and decreased with cat age. The western region of the USA had lower total parasite positivity than other regions for all parasites except Giardia. Cats receiving fecal tests during veterinary wellness visits had only slightly lower parasite positivity than samples from cats during sick clinical visits.ConclusionsThis study showed a larger population of cats are at increased risk of parasitism than commonly believed and coproantigen testing produces more positive test results for the four parasites that antigen can detect than centrifugation of feline fecal samples.

Application of a recombinase polymerase amplification (RPA) assay and pilot field testing for Giardia duodenalis at Lake Albert, Uganda

BackgroundGiardia duodenalis is a gastrointestinal protozoan causing 184 million cases of giardiasis worldwide annually. Detection is by microscopy or coproantigen assays, although sensitivity is often compromised by intermittent shedding of cysts or trophozoites, or operator expertise. Therefore, for enhanced surveillance field-applicable, point-of-care (POC), molecular assays are needed. Our aims were to: (i) optimise the recombinase polymerase amplification (RPA) assay for the isothermal amplification of the G. duodenalis β-giardin gene from trophozoites and cysts, using published primer and probes; and (ii) perform a pilot field validation of RPA at a field station in a resource-poor setting, on DNA extracted from stool samples from schoolchildren in villages around Lake Albert, Uganda. Results were compared to an established laboratory small subunit ribosomal RNA (SSU rDNA) qPCR assay with additional testing using a qPCR targeting the triose phosphate isomerase (tpi) DNA regions that can distinguish G. duodenalis of two different assemblages (A and B), which are human-specific.ResultsInitial optimisation resulted in the successful amplification of predicted RPA products from G. duodenalis-purified gDNA, producing a double-labelled amplicon detected using lateral flow strips. In the field setting, of 129 stool samples, 49 (37.9%) were positive using the Giardia/Cryptosporidium QuikChek coproantigen test; however, the RPA assay when conducted in the field was positive for a single stool sample. Subsequent molecular screening in the laboratory on a subset (n = 73) of the samples demonstrated better results with 21 (28.8%) RPA positive. The SSU rDNA qPCR assay resulted in 30/129 (23.3%) positive samples; 18 out of 73 (24.7%) were assemblage typed (9 assemblage A; 5 assemblage B; and 4 mixed A+B). Compared with the SSU rDNA qPCR, QuikChek was more sensitive than RPA (85.7 vs 61.9%), but with similar specificities (80.8 vs 84.6%). In comparison to QuikChek, RPA had 46.4% sensitivity and 82.2% specificity.ConclusionsTo the best of our knowledge, this is the first in-field and comparative laboratory validation of RPA for giardiasis in low resource settings. Further refinement and technology transfer, specifically in relation to stool sample preparation, will be needed to implement this assay in the field, which could assist better detection of asymptomatic Giardia infections.

Helicobacter pylori infection and gastroduodenal lesions in patients with systemic lupus erythematosus.

The aim of this study was to determine the frequency of Helicobacter pylori in SLE patients and to compare clinical characteristics and gastroduodenal lesions in patients with and without H. pylori infection. Adult SLE patients were selected and subjected to endoscopy. Gastroduodenal lesions were examined by endoscopy and biopsy (antrum and corpus). Biopsies were evaluated by hematoxylin and eosin and Giemsa staining. Immunochromatographic membrane-based assay using amplification was used to test for H. pylori antigen (coproantigen) in stool samples in all participants. Clinical characteristics and gastroduodenal lesions were compared between patients with and without H. pylori infection. A total of 118 SLE patients were included (mean age 44.7 ± 11.7 years, mean disease duration 11.6 ± 6.0 years), of whom 101 (85.6%) were receiving non-steroidal anti-inflammatory drugs (NSAIDs). The coproantigen test was positive in 32 (27.1%) patients. H. pylori was present in twenty six patients (22.0%) in the gastric biopsy. The frequency of gastric erosions and gastric ulcers were 55.1% and 0.8%, respectively. Gastric erosions were less frequent in SLE patients with H. pylori infection than those without H. pylori (43.5.7% vs. 62.5%; p = 0.04). The age, disease duration, disease activity, chronic damage, gastroprotective drugs, and immunosuppressive therapy did not differ between the two groups. We found a high frequency of H. pylori infection in SLE patients. The severity of SLE and reception of gastroprotective therapy do not seem to be related to H. pylori infection. Immunosuppressive therapy may not be protective against H. pylori infection in SLE patients.Key Points• In patients with systemic lupus erythematosus (SLE), the frequency of Helicobacter pylori infection was 39% and gastric erosions were frequent.• Disease activity, chronic damage, gastroprotective drugs, and immunosuppressive therapy may not affect the prevalence of H. pylori infection in SLE patients.

Molecular typing of Cryptosporidium in Israel.

Cryptosporidium is a protozoan parasite associated with gastrointestinal illness. In immune-compromised individuals, the infection may become life-threatening. Cryptosporidiosis is a mandatory-reported disease but little was known about its prevalence and associated morbidity in Israel. Currently, laboratory diagnosis is based on microscopy or copro-antigen tests and the disease is underreported. Molecular assays, which are more sensitive and specific, are now increasingly used for identification and screening. Here, the molecular epidemiology of cryptosporidiosis is explored for the first time. Samples from 33 patients infected during an outbreak of 146 laboratory confirmed cases that occurred in Haifa and Western Galilee in 2015 were genotyped, as well as samples from 36 patients sporadically infected during 2014–2018 in different regions. The results suggest that Cryptosporidium subtypes found in Israel are more similar to those reported in the neighboring countries Jordan and Egypt than in European countries. C. hominis was the predominant species in the center and the north of Israel, implicating human-to-human transmission. C. hominis IeA11G3T3 was the most prevalent subtype contributing to morbidity.

Coproantigen versus microscopic examination for diagnosis of Gardia Lamblia and Entameba Histolytica infection in children in Banha Teaching Hospital

Background: Giardia lamblia (G. lamblia) and Entamoeba histolytica (E. histolytica) are of the most common protozoan enteric pathogen in humans. Their laboratory diagnosis mainly consists of direct microscopic examination of stool specimen for trophozoites and/or cysts. However, due to intermittent fecal excretion of the parasite, the patient may be misdiagnosed and infecting others. Recently, immunological testing of stool (coproantigen) for E histolytica G lamblia has been reported as a more sensitive mean for their diagnosis. Aim of Work: Evaluation of the efficacy of coproantigen detection by ELISA technique in comparison to direct microscopical examination for diagnosis of Entamoeba histolytica and/or Giardia lamblia. Patients and Methods: One hundred ten children were included in this simple comparative study. They were divided into three groups: - Group (1) included 40 children with dysentery their stools were examined for E. histolytica, - Group (2) included 40 children with abdominal complaints their stools were examined for G. lamblia, - Group (3) included 30 normal children their stools were examined for detection of both E histolytica and G. lamblia Results: Group (1) by microscopic examination 35% were positive for trophozoites and/or cysts and 65% were negative while by Coproantigen test 47.5% were positive and 52.5%were negative[sensitivity 73%, specificity 100% and NPV 80.7%]. Group (2) by microscopic examination 27.5%were positive for trophozoites and/or cysts and 72.5% were negative while by Coproantigen test 37.5% were positive and 62.5% were negative [sensitivity 73.3%, specificity 100%,PPV 100% and NPV 86.2%] Group (3) For E. histolytica :by microscopic examination 30% were positive for trophozoites and/or cysts and 70% were negative while by Coproantigen test 23.3% were positive and 76.7% were negative[sensitivity 100%, specificity 91.6% ,PPV 77.7% and NPV 100%] For G. lamblia: by microscopic examination 16.6% were positive for trophozoites and/or cysts and 83.4% were negative (while by Coproantigen test 26.6% were positive and 73.4% were negative [sensitivity 62.5%, specificity 100% , PPV 100% and NPV 89.2%]. Conclusion: Coproantigen ELISA technique is a rapid and effective method with high sensitivity and specificity for diagnosis of E. histolytica and G. lamblia