Carbon Immunoassay Research Articles

Quantitative analysis of TNF-α, IL-4, and IL-10 expression, nitric oxide response, and apoptosis in Encephalitozoon cuniculi–infected rabbits

The expression of tumor necrosis factor (TNF) -α, interleukin (IL) −4 and IL-10, as well as apoptosis and nitric oxide (NO) levels were measured in the brain and kidneys of immunocompetent and immunosuppressed New Zealand White rabbits infected with Encephalitozoon cuniculi. All of the animals had clinical signs histopathological lesions compatible with encephalitozoonosis and were E. cuniculi-positive by using a carbon immunoassay test. Encephalitozoon cuniculi infection promoted the expression of TNF-α and NO production in the kidneys of infected rabbits, and a synergic effect was observed in animal treated with dexamethasone. The IL-4 expression was similar in the brain and kidneys of infected rabbits, regardless of their immunologic status. The IL-10 mRNA expression in the brain of infected immunosuppressed rabbits was elevated when compared with positive controls. Apoptosis of granuloma mononuclear-like cells was detected in immunocompetent E. cuniculi–infected rabbits, but it was more evident in infected-immunosuppressed animals. Nitric oxide levels were elevated both in immunocompetent and immunosuppressed infected animals, but it was more apparent in the kidneys. These data suggest that modulation of the immune response by E. cuniculi could contribute to the survival of the parasite within phagocytic cells in granulomas via an as yet undetermined mechanism.

Serological Investigation on Encephalitozoon cuniculi in pet Rabbits in North-Central Italy

This study determined the seroprevalence of Encephalitozoon cuniculi in pet rabbits from north-central Italy and correlated it with data (age and clinical signs) obtained from private veterinary practitioners. During the period 2007 to 2010, 826 pet rabbit sera were collected and tested using carbon immunoassay. In total, 310 out of 826 rabbits (37.53%, 95% CI: 34.23 to 40.94) displayed clinical signs consistent with infection, whereas the remaining 516 animals presented for wellness examination or to be vaccinated. The results were analyzed according to age, serological status, and the presence or absence of clinical signs. Seropositive results were observed in 59.56% (95% CI: 56.12 to 62.92) of the examined animals. Seroprevalence in the clinically ill rabbits was 70.65% (95% CI: 65.18 to 75.59), much higher than in healthy animals, which showed a prevalence of 52.91% (95% CI: 48.5 to 57.27). Clinical signs were classified into 4 groups: nonvestibular neurologic signs, vestibular signs (including head tilt), renal failure, and ocular lesions. Odds of animals presenting with clinical disease signs were higher in seropositive rabbits (odds ratio = 2.14, 95% CI: 1.59 to 2.9), and this association was particularly true for vestibular signs (odds ratio = 4.1, 95% CI: 2.48 to 7.13). The prevalence values were different with regard to age, with a peak in 3- to 4-year-old animals. Average titers in both healthy and clinically ill rabbits at various ages were different; rabbits with overt disease signs had higher titers when very young (<3 months) and from 1 to 4 years of age. In conclusion, this study provides more precise indications of the prevalence of E. cuniculi in pet rabbits in Italy, underlying growing interest and concern among veterinarians and pet owners regarding this organism.

Encephalitozoon cuniculi in rabbits in Germany: prevalence and sensitivity of antibody testing

The aim of this study was to investigate the prevalence of Encephalitozoon cuniculi antibodies in healthy and diseased rabbits in Germany. Age and gender dependencies were taken into consideration. The...

Serological survey of Encephalitozoon cuniculi infection in commercially reared rabbit does in Northern Italy

The aim of the study was to carry out a serological survey of Encephalitozoon cuniculi in commercially reared rabbit does (Oryctolagus cuniculi) in Veneto region (Northern Italy). Two hundred and sixty blood samples from 13 farms were examined by a carbon immunoassay (CIA test) to detect the presence of antibodies anti-E. cuniculi. All sampled rabbit does were clinically healthy. Seropositivity against E. cuniculi was found in 196/260 (75.4%) sera and in all the sampled farms (100%). Logistic regression analysis showed that the size of the farm had no statistically significant effect on E. cuniculi positivity; whereas rabbits of the hybrid X showed a higher seropositivity (p<0.01) than rabbits belonging to other commercial breeds. Moreover, the age seemed to influence the seropositivity (p<0.05). This serological survey showed a high prevalence of E. cuniculi infection suggesting that this parasite may be endemic in industrial rabbitries in Northern Italy.

Serological survey for antibodies to Encephalitozoon cuniculi in rabbits in Taiwan

Encephalitozoon cuniculi (E. cuniculi) is a microsporidian parasite commonly found in rabbits that can infect humans, causing encephalitozoonosis. Our laboratory recently confirmed the first case of encephalitozoonosis in a rabbit in Taiwan; the prevalence of encephalitozoonosis is not well documented, even when many clinics suspect pet rabbits as being infected. This study surveys the seropositivity of E. cuniculi using carbon immunoassay (CIA) and enzyme-linked immunosorbent assay (ELISA). Serological examination of 171 rabbits using CIA and ELISA showed that 63.2% (108/171) and 67.8% (116/171) were seropositive against E. cuniculi, respectively. Thirteen of the 14 rabbits (92.9%) with neurological symptoms were seropositive. Except for gender, health status and location had a significant effect on E. cuniculi seropositivity (p<0.05). Adult rabbits aged older than 4 months exhibited significantly higher seropositivity for E. cuniculi than young rabbits (p<0.05). In conclusion, this study shows that E. cuniculi is present and widespread among healthy rabbits in Taiwan. Therefore, the fields of veterinary and human medicine in Taiwan should be aware of this zoonotic issue and the resulting public health concern of encephalitozoonosis.

Serological survey of Encephalitozoon cuniculi in farm rabbits in Italy

Rabbit sera ( n = 1600) from 40 commercial farms were submitted to a serological screening for Encephalitozoon cuniculi by an enzyme-linked immunosorbent assay (ELISA) and a carbon immunoassay (CIA test). Antibodies anti- Encephalitozoon cuniculi were found in 505/1600 (31.6%) sera analysed, and all the farms (100%) resulted positive. Rabbits older than 4 months showed a significantly higher seropositivity for E. cuniculi (chi-squared test: p < 0.0001) than rabbits under 4 months, E. cuniculi sero-prevalence showed an increasing trend in rabbits within the farm along with the increase in the “number of rabbits on the farm”; however, this trend was not significant (Spearman’s correlation: p = 0.073). The findings of the present study confirm that rabbit is the main reservoir of E. cuniculi; they are of epidemiological relevance and immediate public health importance because of the recognized infectivity in humans by the microsporidium.

Serological Survey for Antibodies to Encephalitozoon cuniculi in Pet Rabbits in Italy

Pet rabbits (n = 125) from Southern Italy were submitted to a serological screening for Encephalitozoon cuniculi, using an enzyme-linked immunosorbent assay (ELISA) and a carbon immunoassay (CIA). Seventy-eight examined rabbits showed clinical signs suggestive of encephalitozoonosis (head tilt, ataxia, paralysis, cataracts, uveitis, polyuria and polydipsia), whereas 47 were healthy rabbits. Antibodies anti-E. cuniculi were found in 84/125 (67.2%) sera analysed. The results of the chi-squared test showed that sex and health status had no significant effect (P > 0.05) on E. cuniculi seropositivity; however, rabbits older than 4 months had a seropositivity for E. cuniculi significantly (P < 0.05) higher than that of rabbits aged up to 4 months. The results of the present survey reinforce the assumption that rabbit may be indicated as the main reservoir of E. cuniculi; therefore, routine screening examinations in pet rabbits are strongly advised considering the zoonotic potential of this parasite.

Immunohistochemical identification of Encephalitozoon cuniculi in phacoclastic uveitis in four rabbits

Encephalitozoon cuniculi is a microsporidium with a wide range of mammalian hosts. In rabbits it can be responsible for cataract and lens-induced uveitis (LIU). The aim of this study was to provide specific immunohistochemical demonstration and localization of E. cuniculi within the eye, in rabbits with LIU. Four rabbits were presented with a white mass in the eye and iris discoloration. Complete ophthalmic examinations were performed and a presumptive diagnosis of LIU was made in all cases. Initial therapy with a topical steroid, atropine and systemic enrofloxacin was instituted while serologic (IFA or ICA tests) and cytologic lab results were pending. The final outcome in all cases was enucleation. Routine histology and immunohistochemistry (ABC method) with an antiserum anti-Encephalitozoon cuniculi were performed. Indirect immunofluorescence performed on one rabbit serum expressed a titer of 1 : 32; carbon immunoassay on the serum of the other three rabbits expressed a titer of 1 : 5120 in one, and a titer of 1 : 2560 in the other two cases. Histologically, an intraocular, locally extensive pyogranulomatous infiltration that partially filled the posterior chamber, encasing a wide anterior lens capsule break, was detected in all cases. Immunohistochemically, spores reacting with anti-Encephalitozoon cuniculi antiserum were present in all specimens, occasionally within macrophages and lens epithelial cells. Detection of E. cuniculi in rabbits with phacoclastic uveitis has been investigated in the past with different methods. Based on our results, we suggest that immunohistochemistry should be regarded as a useful tool both for specific demonstration of E. cuniculi and for its localization within tissues.

Humoral immune response in adult blue foxes ( Alopex lagopus) after oral infection with Encephalitozoon cuniculi spores

Encephalitozoon cuniculi causes severe diseases in blue fox puppies. When pregnant vixens are infected, parasites are transmitted over the placenta to the unborn that subsequently develop encephalitozoonosis. Adult foxes themselves do not have signs of disease, but show antibody titres to E. cuniculi. The purpose of the present study was to gain information on the immune response in adult foxes after experimental infection. Sixteen foxes were infected orally with E. cuniculi spores, eight of them twice and 28 days apart. The two groups of animals showed elevated serological values in both the carbon immunoassay and in the ELISA. Elevated serological levels were recorded up to 1 year after the infection took place. The control group ( n=8) remained serologically negative throughout the trial. The results of the study showed that blue foxes could be seropositive for at least a year after oral infection with E. cuniculi.

Survey for Encephalitozoon cuniculi in Arctic Foxes (Alopex lagopus) in Greenland

Wild arctic foxes (Alopex lagopus) from Greenland were tested for antibodies to Encephalitozoon cuniculi with an enzyme-linked immunosorbent assay and a carbon immunoassay. Of 230 tested foxes none was seropositive. This finding contrasts with observations from other arctic areas and absence of rodents in the diet of these arctic foxes is the most likely explanation for absence of E. cuniculi.

Determination of Toxoplasma gondii in several organs of cattle by carbon immunoassay (CIA) testing

Samples of muscle, heart, liver and lung were tested for the presence of Toxoplasma gondii using mice as a biologic detector and the carbon immunoassay (CIA) for immunological diagnosis. In positive tissues the percentage of samples positive ranged from 10 to 50% which indicates the importance of cattle in the epidemiology of Toxoplasma in Costa Rica. We discuss these findings in relation to the transmission of the parasite in Costa Rica.

Specificity of the carbon immunoassay (CIA) test for the diagnosis of Toxoplasma infections

A rapid and low cost procedure, the carbon immunoassay (CIA) test, was evaluated for the diagnosis of Toxoplasma gondii infections. Using a closely related parasite ( Besnoitia jellisoni) as antigen, and homologous or heterologous immune sera, it was demonstrated by light and electron microscopy that CIA is a very reliable and specific test. As it is neither expensive nor time-consuming, it can be recommended for general and routine laboratory use.

The incidence of Encephalitozoon cuniculi in a commercial barrier-maintained rabbit breeding colony

Between 1982 and 1987 sera from 4952 New Zealand White rabbits (Oryctolagus cuniculus) obtained from a single commercial supplier were tested for the presence of antibodies to Encephalitozoon cuniculi. A commercially available carbon immunoassay test kit was used. Initially 32.9% of the rabbits were seropositive with the number progressively decreasing to 2.3% by 1987. The reason for the significant decline in the incidence of infection was most likely due to a selection process for breeding stock instituted by the supplier based upon productivity, posture and weight of each animal.

Humoral antibody response of rabbits to experimental infection with Encephalitozoon cuniculi

Six female rabbits ( Oryctolagus cuniculus) were experimentally infected intravenously with approximately 1.5 × 10 7 live spores of Encephalitozoon cuniculi. Head tilt was observed as the single clinical sign in only one of the six animals. Antibody response was registered over 68 days postinfection using the indirect immunofluorescence test (IFT) for IgM and IgG, and the carbon immunoassay (CIA). IgG titers reached a level of 160–2560 after a latent phase of 13–28 days, followed by a 2–4 week relatively steep increase. The IgM seroconversion was faster than that of IgG and occurred at the beginning of the antibody response. Thus, simultaneous detection of both IgM and IgG allowed the infection to be identified as recent. Long, short, and episodic antibody responses could be distinguished: the IgG titer continued to increase on Day 68 in one animal (long response) and began to decrease between Days 45 and 63 in three other animals (short response). In two additional animals the seroconversion was very short, occuring between Days 13 and 41, and 28 and 52, respectively (episodic response). The CIA proved to be specific, reliable, and simple to perform; titers were slightly higher than in the IFT. Parasite pseudocysts were detected scattered throughout the brain on Day 68 in four of the six rabbits. The persistence of antigen in the brain did not correlate with antibody response, which in most cases was shorter.

Application of an indirect carbon immunoassay (CIA) for the rapid diagnosis of antibody to [formula omitted] in sheep

Carbon immunoassay (CIA), a novel indirect rapid test for Toxoplasma antibody in sheep, was compared with indirect fluorescent antibody assay (IFA). CIA relies on the adherence of carbon particles of India-ink to rabbit immunoglobulin G. Carbon labelled anti-sheep rabbit IgG was used for the detection of sheep antibody when attached to tachyzoites of Toxoplasma gondii . The result was read in an ordinary light microscope and there was a clearcut difference between negative and positive reactions. Out of a total of 97 sheep sera tested, 15 sera were negative in both tests and 12 were negative in CIA but showed low positive titres in IFA. The remaining 70 sera were positive in both tests but the titres were usually about 3 dilution steps lower when investigated with CIA as compared to IFA.

A novel simple immunoassay for rapid detection of human IgG antibodies to toxoplasma gondii

• The carbon immunoassay (CIA) is a direct serological test relying on a specific reaction between the carbon particles of India ink and rabbit immunoglobulin G. On addition of the India ink reagent, particulate antigens may be visualized in the light microscope after rabbit antibodies have reacted with them. Serum, India ink reagent and antigen may be mixed together in a single step and the result read within 5 min. The applicability of CIA can be expanded to include human sera by pretreatment of India ink with staphylococcal protein A. • Sera from 50 suspected toxoplasmosis patients were examined by immunofluorescence (IF), the dye test of Sabin-Feldman (SF) and CIA. There was good correlation between results by the CIA and IF. Sera with low SF titres were usually negative by IF and negative or weakly positive by CIA. With sera positive by all 3 tests, CIA and IF gave similar titres, but SF titres were generally higher. • CIA provides a simple inexpensive and rapid laboratory test for diagnosis of toxoplasmosis.