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BACKGROUND. Intravenous Ig (IVIg), plasma exchange, and immunoadsorption are frequently used in the management of severe autoimmune diseases mediated by pathogenic IgG autoantibodies. These approaches modulating IgG levels can, however, be associated with some severe adverse reactions and a substantial burden to patients. Targeting the neonatal Fc receptor (FcRn) presents an innovative and potentially more effective, safer, and more convenient alternative for clearing pathogenic IgGs. METHODS. A randomized, double-blind, placebo-controlled first-in-human study was conducted in 62 healthy volunteers to explore single and multiple ascending intravenous doses of the FcRn antagonist efgartigimod. The study objectives were to assess safety, tolerability, pharmacokinetics, pharmacodynamics, and immunogenicity. The findings of this study were compared with the pharmacodynamics profile elicited by efgartigimod in cynomolgus monkeys. RESULTS. Efgartigimod treatment resulted in a rapid and specific clearance of serum IgG levels in both cynomolgus monkeys and healthy volunteers. In humans, single administration of efgartigimod reduced IgG levels up to 50%, while multiple dosing further lowered IgGs on average by 75% of baseline levels. Approximately 8 weeks following the last administration, IgG levels returned to baseline. Efgartigimod did not alter the homeostasis of albumin or Igs other than IgG, and no serious adverse events related to efgartigimod infusion were observed. CONCLUSION. Antagonizing FcRn using efgartigimod is safe and results in a specific, profound, and sustained reduction of serum IgG levels. These results warrant further evaluation of this therapeutic approach in IgG-driven autoimmune diseases. TRIAL REGISTRATION. Clinicaltrials.gov {"type":"clinical-trial","attrs":{"text":"NCT03457649","term_id":"NCT03457649"}}NCT03457649. FUNDING. argenx BVBA.

Encephalitozoon intestinalis (Septata intestinalis) is the second most prevalent microsporidian species infecting humans, but it has not been described in other animal species. This investigation examined 10 domestic animal stool samples (8 mammalian, 2 avian) containing spores detected by anti-Encephalitozoon monoclonal antibody immunofluorescence (FA). The presence of E. intestinalis but not Encephalitozoon hellem or Encephalitozoon cuniculi was confirmed in 6 of 8 mammalian stool samples by species-specific FA and polymerase chain reaction. Clusters of spores inside epithelial cells were observed in feces of five mammals (donkey, dog, pig, cow, and goat) using "quick-hot" Gram-chromotrope stain. None of the 10 samples reacted with anti-E. hellem or anti-E. cuniculi sera, nor were they amplified with species-specific primers for E. hellem and E. cuniculi. To our knowledge, this is the first identification of E. intestinalis in animals other than humans. The data shown herein suggest the possibility that E. intestinalis infection may be zoonotic in origin.

The term microsporidia is used to describe several species of opportunistic protozoan parasites. Encephalitozoon intestinalis and Enterocytozoon bieneusi have been found in stools of more than 40% of AIDS patients with diarrhea. Diagnosis of infection with these small protozoans has been difficult, and until recently their occurrence has not been well documented. Formalin is widely used to preserve clinical specimens, but due to the nature of the fixation process, subsequent analysis, especially analysis by the PCR, is difficult. This study evaluated methods used to prepare formalin-fixed fecal specimens for PCR amplification of microsporidial DNA. Two methods were devised to allow PCR detection and subsequent identification of microsporidia in formalin-fixed fecal specimens to the species level. One method involved immunomagnetic separation to concentrate microsporidial spores from fecal specimens. In the second method Chelex resin (Bio-Rad, Hercules, Calif.) was used to remove inhibitory substances, followed by a DNA concentration step. Both methods resulted in reproducible, confirmed detection of microsporidia in formalinized fecal specimens and subsequent species determination by PCR sequencing. The detection sensitivity was two in vitro culture-derived spores ( Encephalitozoon intestinalis ) for the direct PCR. The reproducible detection sensitivity for DNA amplification from formalin-fixed fecal samples was 200 spores for either the Chelex method or the immunomagnetic bead separation method. Thus, we developed two methods for rapid, inexpensive detection of microsporidial spores in formalin-fixed fecal specimens.

Using an indirect immunofluorescence assay on stool samples, we found a 6.4% prevalence of cryptosporidiosis among 403 children less than five years of age with acute diarrhea in Mexico City over a one-year period. The prevalence was highest (11.4%) during the rainy summer months. Most Cryptosporidium parvum cases occurred in infants less than one year of age. Cryptosporidium parvum was more common in malnourished children (P < 0.05) and in nonbreast-fed infants less than six months of age (P < 0.01). Neither dwelling characteristics nor the presence of domestic animals at home were associated with C. parvum infection. Enteropathogenic bacteria were found in 26.8% of the children; Escherichia coli, Salmonella, and Shigella being the most frequently identified. None of 100 serum samples tested showed antibodies against human immunodeficiency virus. When compared with immunofluorescence, the acid-fast technique showed a sensitivity of 76.9% and a specificity of 98.9%, with a predictive positive value of 83.3%. It was concluded that 1) monoclonal antibody-based immunofluorescence is a better diagnostic tool than the acid-fast technique, 2) the prevalence of cryptosporidiosis in this population was similar to that of other developing countries, 3) clinical manifestations were nonspecific, and 4) C. parvum was more common in malnourished children and in nonbreast-fed infants.

Infections with Encephalitozoon (phylum Microspora) protozoa in immunodeficient patients have emerged worldwide. The prevalence of infections due to Encephalitozoon species in nonclinical populations remains unclear. We conducted a cross-sectional survey of two rural highland villages in Mexico by using monoclonal antibody 3B6 to Encephalitozoon in immunofluorescence to assess the prevalence of Encephalitozoon spores in human stools. We found that 20 (7.84%) of the 255 subjects were positive and that 15 (21.4%) of the 70 households had at least one member who was positive. These results suggest that Encephalitozoon species, most likely Encephalitozoon intestinalis, may be commonly present in community settings.

Differences in susceptibility to persistent cryptosporidial infection between two strains of adult athymic nude mice prompted us to investigate the immune mechanism(s) that may control resistance to infection in these T-cell-deficient mice. We studied fecal oocyst shedding, serum and fecal parasite-specific antibody responses, and fecal immunoglobulin levels in athymic C57BL/6J nude and athymic BALB/cJ nude mice following oral inoculation with Cryptosporidium parvum oocysts at 8 to 9 weeks of age. C57BL/6J nude mice had significantly higher fecal parasite-specific immunoglobulin A (IgA) (days 27, 31, 35, and 42 postinoculation) and IgM (days 10, 17, 24, 28, 31, 38, 42, and 48 postinoculation) levels than BALB/cJ nude mice ( P < 0.05) and significantly higher serum parasite-specific IgA levels at 63 days postinoculation ( P < 0.03). Moreover, C57BL/6J nude mice shed significantly fewer C. parvum oocysts than BALB/cJ nude mice from days 52 to 63 postinoculation ( P < 0.05). In contrast, BALB/cJ nude mice had higher levels of non-parasite-specific IgA (days 38 to 63 postinoculation) and IgM (days 24, 35, 38, and 52 postinoculation) than C57BL/6J nude mice in feces ( P < 0.05). These data suggest that parasite-specific fecal antibodies may be associated with resistance to C. parvum in C57BL/6J nude mice.

Cryptosporidium parvum is an important diarrhea-causing protozoan parasite of immunocompetent and immunocompromised hosts. Immunoglobulin A (IgA) has been implicated in resistance to mucosal infections with bacteria, viruses, and parasites, but little is known about the role of IgA in the control of C. parvum infection. We assessed the role of IgA during C. parvum infection in neonatal mice. IgA-secreting hybridomas were developed by using Peyer’s patch lymphocytes from BALB/c mice which had been orally inoculated with viable C. parvum oocysts. Six monoclonal antibodies (MAbs) were selected for further study based on indirect immunofluorescence assay reactivity with sporozoite and merozoite pellicles and the antigen (Ag) deposited on glass substrate by gliding sporozoites. Each MAb was secreted in dimeric form and recognized a 23-kDa sporozoite Ag in Western immunoblots. The Ag recognized comigrated in sodium dodecyl sulfate-polyacrylamide gel electrophoresis with P23, a previously defined neutralization-sensitive zoite pellicle Ag. MAbs were evaluated for prophylactic or therapeutic efficacy against C. parvum , singly and in combinations, in neonatal BALB/c mice. A combination of two MAbs given prophylactically prior to and 12 h following oocyst challenge reduced the number of intestinal parasites scored histologically by 21.1% compared to the numbers in mice given an isotype-matched control MAb ( P < 0.01). Individual MAbs given therapeutically in nine doses over a 96-h period following oocyst challenge increased efficacy against C. parvum infection. Four MAbs given therapeutically each reduced intestinal infection 34.4 to 42.2% compared to isotype-matched control MAb-treated mice ( P < 0.05). One MAb reduced infection 63.3 and 72.7% in replicate experiments compared to isotype-matched control MAb-treated mice ( P < 0.0001). We conclude that IgA MAbs directed to neutralization-sensitive P23 epitopes may have utility in passive immunization against murine C. parvum infection.