Indirect Immunofluorescent Antibody Research Articles

Evaluation of the Accuracy of Estimated Endpoint Titer of NOVA View in Indirect Immunofluorescent Antinuclear Antibody Testing

For antinuclear antibody (ANA) screening, the gold standard method is an indirect immunofluorescence assay (IIFA) using HEp-2 cells, and a serial dilution test is needed to determine the endpoint titer. We aimed to evaluate the accuracy of the estimated endpoint titer (eEPT) by the NOVA View system, by comparing it with the EPT by the serial dilution method (dEPT). The endpoint titers of a total of 1518 ANA positive cases with five major patterns including speckled, homogeneous, centromere, nucleolar, and nuclear dots patterns were determined using both the estimation function and the serial dilution method by the NOVA View system. A significant correlation between the light intensity unit (LIU) values and dEPTs was identified in all five patterns with high ρ values, ranging from 0.666 to 0.832. However, the overall exact match rate between dEPT and eEPT was 22.1% (336/1518), with the ±one-titer match rate being highest in the centromere pattern (62.8%, 81/129), and lowest in the homogeneous pattern (37.6%, 200/532). This suggests that while LIU values correlate well with dEPT, there are discrepancies in numerical agreement. Most cases that did not show an exact match, showed one-to-three-titer overestimations by eEPT. Therefore, adjusting eEPT downward significantly improved the concordance rates with dEPTs. Further investigation for an appropriate cutoff of LIU values for determining eEPT should be performed for clinical application and contribution to the standardization of the ANA titer.

Assessing serum anti-nuclear antibodies HEp-2 patterns in synucleinopathies

This study investigates the presence of antinuclear antibodies (ANA) in three primary synucleinopathies – Parkinson’s disease (PD), multiple system atrophy (MSA), and dementia with Lewy bodies (DLB), compared to healthy controls. Autoinflammatory disorders typically involve the immune system mistakenly attacking the body’s own cells and start producing ANA. There is an increasing body of evidence that immune-mediated inflammation is a pathological feature linked to synucleinopathies. To investigate whether this could be autoimmune mediated we analyzed for ANA in the plasma of 25 MSA, 25 PD, and 17 DLB patients, along with 25 healthy controls, using the ANA HEp-2 indirect immunofluorescence antibody assay (ANA HEp-2 IFA). Contrary to initial expectations, results showed ANA HEp-2 positivity in 12% of PD, 8% of MSA patients, 18% of DLB patients, and 17% of healthy controls, indicating no increased prevalence of ANA in synucleinopathies compared to age-matched healthy individuals. Various ANA HEp-2 patterns were identified, but no specific pattern was associated with individual synucleinopathies. We conclude hereby that synucleinopathies are not associated with detectable presence of ANA in plasma.

Serological Screening and Risk Factors Associated with Leishmania infantum Positivity in Newly Diagnosed HIV Patients in Greece.

A serological screening was conducted to detect IgG antibodies against Leishmania infantum (L. infantum) in newly diagnosed human immunodeficiency virus (HIV) patients in Greece. The study also examined potential risk factors and the agreement of commercially available serological methods. IgG antibodies against L. infantum were detected using enzyme-linked immunosorbent assay (ELISA), indirect immunofluorescence antibody test (IFAT), and Western blot (WB). Out of 155 samples, 14 (9.0%) tested positive for IgG antibodies against L. infantum using at least two methods. Statistical analysis showed substantial agreement between WB and IFAT methods (Cohen's kappa = 0.75) but moderate overall agreement among the three methods (Fleiss' kappa = 0.42). Additionally, HIV+ intravenous drug users faced 3.55 times (p = 0.025) higher risk of testing positive for L. infantum IgG, positing that anthroponotic transmission between these patients is a plausible hypothesis based on existing literature. Non-invasive and cost-effective techniques are preferred to detect asymptomatic infections, and leishmaniasis screening should be conducted immediately after HIV diagnosis in endemic regions to enable prophylactic treatment for leishmaniasis in addition to antiretroviral therapy. To maximize sensitivity, performing at least two different serological methods for each patient is recommended.

Seroprevalence of Toxoplasma gondii among HIV Positive Patients under Surveillance in Greek Infectious Disease Units: A Screening Study with Comparative Evaluation of Serological Methods.

This study aims to screen for IgG antibodies against Toxoplasma gondii (T. gondii) in the sera of 155 newly diagnosed Human Immunodeficiency Virus (HIV) positive patients under surveillance in Greek Infectious Disease Units. Additionally, risk factors based on patient demographics were examined, and a comparative evaluation of commercially available serological methods was conducted. Three methods were employed to detect IgG antibodies against T. gondii: Enzyme-Linked Immunosorbent Assay (ELISA), Indirect Immunofluorescence Antibody Test (IFAT), and Western Blot (WB), which was used as a reference here. Forty-nine sera samples were true-positive for IgG antibodies against T. gondii, resulting in a 31.61% positivity rate, and the immunoassay test statistical reliability analysis resulted in higher IFAT accuracy (90.97%) compared to ELISA (76.26%). Furthermore, statistical analysis of demographic and immunological data included in the study placed female and foreign/non-Greek individuals at 2.24 (p = 0.0009) and 2.34 (p = 0.0006) times higher risk of positive T. gondii IgG testing compared to their male and Greek counterparts, respectively. Our findings on positivity rates and comparative serology underscore the importance of early and suitable screening measures for newly diagnosed HIV+ patients to mitigate the life-threatening outcomes that may arise from a potential subsequent T. gondii activation.

Detection of Anaplasma spp. and Ehrlichia spp. in dogs from a veterinary teaching hospital in Italy: a retrospective study 2012–2020

Anaplasma phagocytophilum, Anaplasma platys and Ehrlichia canis, responsible of diseases in dogs, are tick-borne pathogens with a proven or potential zoonotic role that have shown increasing prevalence worldwide. The aims of this retrospective study were to assess the frequency of Anaplasma spp. and Ehrlichia spp. exposure in dogs tested in a veterinary teaching hospital in Italy over a 9-year period, to compare the performance of the diagnostic tests used, to evaluate correlations with clinical data, and to genetically analyse the identified bacteria. During the study period, 1322 dogs tested by at least one of the rapid immunoenzymatic test, indirect immunofluorescent antibody test or end-point PCR assay for Anaplasmataceae detection were included. Dogs were tested if they had clinical signs or clinicopathological alteration or risk factors related to infection, and if they were potential blood-donor animals. Ninety-four of 1322 (7.1%) dogs tested positive for at least one pathogen: 53 (4.3%) for A. phagocytophilum, one (0.1%) for A. platys and 63 (4.6%) for E. canis. The number of dogs tested increased and the positivity rate progressively declined over the years. Comparison of tests showed a near-perfect agreement between serological tests and a poor agreement between PCR and indirect assays. A breed predisposition has been highlighted for A. phagocytophilum infection in hunting breed dogs and for E. canis infection in mixed breed dogs. Phylogeny confirmed potential zoonotic implications for A. phagocytophilum and showed no correlation of the identified bacteria with the geographical origin. Our study provides new insights into possible risk factors in dogs and evidenced discordant results between different tests, suggesting that a combination of serological and molecular assays is preferable for a correct diagnosis.

Toxoplasmosis Behind Bars: One Health Approach on Serosurvey Dynamics and Associated Risk Factors for Women Inmates, Correctional Officers, and In-Prison Feral Cats

Brazil holds the third highest general and fifth female incarcerated population worldwide. Despite the incarceration ecosystem that may favor the spreading of zoonotic diseases, particularly when unattended animals are present, no comprehensive study has focused on toxoplasmosis dynamics in such environment. Accordingly, the present study has aimed to serologically assess anti-Toxoplasma gondii (IgG) antibodies by indirect immunofluorescent antibody test in inmates, correctional officers, and feral cats at the Women’s State Penitentiary of Parana, southern Brazil. In overall, 230/506 (45.5%; CI 95%: 41.2–49.8) incarcerated women, 31/91 (34.1%; 95% CI: 25.2–44.3) correctional officers, and 23/39 (59.0%; CI 95%: 43.2–72.9) cats were seropositive to anti-T. gondii antibodies. Logistic regression revealed that seropositivity likelihood increased with consumption of raw meat (p=0.040) and decreased with elementary educational level (p=0.001). No statistical difference was found comparing seropositivity between inmates and correctional officers (p=0.057). As women inmates have been considered among the most vulnerable groups in disease morbidity and mortality, seropositivity observed herein may be directly related to vulnerability and high T. gondii oocyst exposure dispersed in cat feces during incarceration.

Spontaneous splenic rupture associated with scrub typhus: a case report

BackgroundScrub typhus, an acute febrile disease with mild to severe, life-threatening manifestations, potentially presents with a variety of complications, including pneumonia, acute respiratory distress syndrome, cardiac arrhythmias (such as atrial fibrillation), myocarditis, shock, peptic ulcer, gastrointestinal bleeding, meningitis, encephalitis, and renal failure. Of the various complications associated with scrub typhus, splenic rupture has rarely been reported, and its mechanisms are unknown. This study reports a case of scrub typhus-related spontaneous splenic rupture and identifies possible mechanisms through the gross and histopathologic findings.Case presentationA 78-year-old man presented to our emergency room with a 5-day history of fever and skin rash. On physical examination, eschar was observed on the left upper abdominal quadrant. The abdomen was not tender, and there was no history of trauma. The Orientia tsutsugamushi antibody titer using the indirect immunofluorescent antibody test was 1:640. On Day 6 of hospitalization, he complained of sudden-onset left upper abdominal quadrant pain and showed mental changes. His vital signs were a blood pressure of 70/40 mmHg, a heart rate pf 140 beats per min, and a respiratory rate of 20 breaths per min, with a temperature of 36.8 °C. There were no signs of gastrointestinal bleeding, such as hematemesis, melena, or hematochezia. Grey Turner's sign was suspected during an abdominal examination. Portable ultrasonography showed retroperitoneal bleeding, so an emergency exploratory laparotomy was performed, leading to a diagnosis of hemoperitoneum due to splenic rupture and a splenectomy. The patient had been taking oral doxycycline (100 mg twice daily) for 6 days; after surgery, this was discontinued, and intravenous azithromycin (500 mg daily) was administered. No arrhythmia associated with azithromycin was observed. However, renal failure with hemodialysis, persistent hyperbilirubinemia, and multiorgan failure occurred. The patient did not recover and died on the fifty-sixth day of hospitalization.ConclusionsClinicians should consider the possibility of splenic rupture in patients with scrub typhus who display sudden-onset abdominal pain and unstable vital signs. In addition, splenic capsular rupture and extra-capsular hemorrhage are thought to be caused by splenomegaly and capsular distention resulting from red blood cell congestion in the red pulp destroying the splenic sinus.Graphical abstract

Retrospective analysis of the clinical significance of Ro52/TRIM21 antibody and specific antinuclear antibody patterns by indirect immunofluorescence.

To determine the clinical significance of Ro52 protein/tripartite motif-containing 21 antibody and specific antinuclear antibody patterns using indirect immunofluorescence technique. The retrospective study was conducted at the clinical laboratory of the First Affiliated Hospital of Chongqing Medical University, China, and comprised data from January 2017 to December 2021 of patients who underwent antinuclear antibody and anti-extractable nuclear antigen antibody detection. Inpatients with Ro52 antibody-positive status were taken as the cases, while anti-Ro52 negative patients with clear clinical diagnosis were taken as the controls. Data was analysed using SPSS 19. There were 1802 cases and 1211 controls. Positive Ro52 showed significantly greater frequency in patients with primary Sjogren's syndrome, systemic lupus erythematosus, inflammatory myositis, dry eyes and interstitial lung disease (p<0.05). Ro52 antibody showed high positive predictive value for primary Sjogren's syndrome 25(96.15%), systemic lupus erythematosus 259(91.20%), connective tissue disease-associated interstitial lung disease 45(86.67%) and inflammatory myositis 60(86.67%). Antinuclear antibody indirect immunofluorescence patterns most frequently detected were nuclear speckled 128(40.89%) and cytoplasmic speckled 126(40.26%) (p<0.05). Interstitial lung disease was associated with the presence of cytoplasmic speckled antinuclear antibody indirect immunofluorescence pattern 24(19.2%), while tumours 47(36.5%) and hepatitis B 26(20.3%) seemed to be more frequent with nuclear speckled pattern (p<0.05). The simultaneous reactivity extractable nuclear antigen antibodies most frequently detected were antinuclear antibody+Ro52+anti-Sjogren's syndrome A+ 558(33.96%). Ro52 antibody positivity was found to be associated with Sjogren's syndrome, systemic lupus erythematosus, inflammatory myositis, dry eye and interstitial lung disease. The antinuclear antibody immunofluorescence pattern of Ro52 positive was single and primarily granular cytoplasm type. Antinuclear antibody negative and Ro52 positive in the serum of patients also had certain significance in auxiliary disease diagnosis.

Seroprevalence of Toxoplasma gondii in wild boars (Sus scrofa) hunted in Ukraine

Toxoplasma gondii is an important zoonotic parasite worldwide, but it has received limited attention in Ukraine. A seroepidemiological study was conducted and samples from 452 wild boars that had been hunted in 2006–2011 in 23 of the 25 regions of Ukraine were tested to estimate T. gondii seroprevalence. A locally available commercial enzyme-linked immunosorbent assay (ELISA) was used for the investigation. Additionally, we tested 92 of the sera using a widely used commercial multi-species ELISA and an indirect immunofluorescence antibody test (IFAT). With the locally available ELISA, 35 of the 452 wild boars tested positive, yielding a seroprevalence estimate of 7.7% (95% confidence interval 5.5–10.5). The seropositive wild boars originated from eight of the regions. Using the majority criteria, 10/92 samples tested using both ELISAs and the IFAT were considered positive, yielding an estimated seroprevalence of 10.9% within the subset of samples. The highest seroprevalence was observed in wild boars hunted in Luhans'k (30.0%), Odesa (17.7%) and Kharkiv (12.7%). Seroprevalence was higher in older animals (13.3% for age group >12 months and 7.7% for age group ≤12 months). This is the first seroepidemiological study of T. gondii in wild boars in Ukraine. Assuming that seropositivity indicates presence of infectious parasites in the tissues, eating undercooked meat of wild boars hunted in Ukraine could be a potential source of infection to other hosts, including humans.

Comparison of laboratory methods for diagnosis of human visceral leishmaniasis in the Center-West Region of São Paulo State, Brazil

Visceral leishmaniasis (VL) is a neglected disease and represents an important public health problem. The present study aimed to compare the conventional laboratory methods with the molecular method for VL diagnosis in human blood samples. Direct parasitological test of bone marrow aspirate (direct parasitological test), rapid immunochromatographic test (RIT), indirect immunofluorescent antibody test (IFAT) and polymerase chain reaction (PCR) were used to evaluate 26 human samples with clinical suspicion of VL. The positivity rates obtained for direct parasitological test, RIT, IFAT and PCR were 80.8%, 80.8%, 73.1% and 84.6%, respectively. PCR had greater positivity and agreement with the other methodologies. Therefore, the use of PCR in the diagnostic routine should be encouraged to complement the serological results.

The Discrepancy of ANA and Compartment Bead Patterns Suggestive of a Neuropsychiatry Systemic Lupus Erythematosus (NPSLE)

Neuropsychiatric systemic lupus erythematosus (NPSLE) exhibits neurological and psychiatric manifestations in systemic lupus erythematosus (SLE) patients, which NPSLE diagnosis can be challenging for rheumatologists. An Indonesian female, 44 years old, complained of two times seizures with 10-min duration, which during seizures were stiff, eyes rolled up, foaming at the mouth, wet the bed, and fainting afterward. The patient also has a history of SLE and received cyclophosphamide therapy 5 years ago. Her clinical condition showed facial and lingual palsy, with central type on the right. Antinuclear antibody indirect immunofluorescence (ANA IF) positive using cytobead ANA with a homogenous pattern and cytoplasmic speckled titer 1/80. Confirmation beads showed positive of dsDNA only. ANA profile showed positive antinucleosome, antihistone, and AMA-M2, and also increased anticardiolipin antibody that supports the diagnosis of NPSLE. The difference in the pattern of ANA IF with confirmation beads suggests the presence of other autoantibodies in NPSLE.

Five-Year Analysis of Anti-Nuclear Antibody (ANA) and Extractable Nuclear Antigen Antibody (anti-ENA) Immunoblot Test Results

Autoimmune disorders may cause systemic disease in which multiple organs are affected or may also be localized as single-organ involvement. Various ancillary immunological tests used in cases where systemic involvement is observed, and differential diagnosis becomes difficult or non-specific symptoms are observed. For this purpose, investigation of antinuclear antibodies (ANA) and antibodies against extractable nuclear antigens (ENA) in patient serums using various methods is a guide for clinicians. In this study, we aimed to evaluate the effectiveness of ANA and ENA antibody tests by examining the test results, the patients’ demographic data, and the clinics that ordered the tests. In the study, a retrospective review of the results of 37,791 ANA tests studied on serum samples sent to the medical microbiology laboratory of our hospital from various clinics between January 2018 and December 2022 is presented. ANA tests were resulted semi-quantitatively by evaluating the slides prepared using diluted serum samples at a ratio of 1:100 and HEp-2 cells and monkey liver tissue under a fluorescent microscope with the indirect immunofluorescence (IIF) antibody method. Of the patients who requested ANA test, 11,604 (30.7%) were male and 26,187 (69.3%) were female. The mean ages were 43.46 (0-94) and 46.94 (0-96), respectively. Of the samples tested, 18.8% (7,107/37,791) were evaluated as ANA positive. The ANA positivity rate was significantly higher in women (24.8%) than in men (19.7%) (p&lt;0.001). ANA positive samples were re-tested using the anti-ENA immunoblot method. Of the 7,107 ANA-positive patients, 3,809 (53.6%) were found to be anti-ENA positive for at least one specific antibody, while 3,298 (46.4%) were negative. In the anti-ENA profile, antibodies against 11 different antigen types were screened and the most frequently detected antibody was anti-SS-A (22.7%), and this was followed by anti-centromere (12.8%), anti-RNP (10.2%), and anti-dsDNA (9.7%) antibodies. Immunoblot-based anti-ENA tests provide specific information for the diagnosis of autoimmune diseases. In our study, the fact that anti-ENA tests were negative in a significant part of ANA positive patients indicates the inadequacy of the immunoblot test panel and reveals the need for immunoblot tests with wider antibody diversity or any alternative methods.

Identification of specific antigens between Toxoplasma gondii and Neospora caninum and application of potential diagnostic antigen TgGRA54.

Toxoplasma gondii is a zoonotic parasite that is very common in livestock. Meat products from livestock infected with T. gondii are one of the important transmission routes of toxoplasmosis. Rapid and reliable diagnosis is a prerequisite for the prevention and control of toxoplasmosis. Neospora caninum and T. gondii are similar in morphology and life history, and there are a large number of cross antigens between them, making clinical diagnosis of toxoplasmosis more difficult. In this study, immunoprecipitation-mass spectrometry (IP-MS) was used to screen for T. gondii-specific antigens, and the specific antigen was cloned and expressed in Escherichia coli. The specific antigen was then used to establish an indirect ELISA diagnostic method. A total of 241 specific antigens of T. gondii and 662 cross antigens between T. gondii and N. caninum were screened by IP-MS. Through bioinformatics analysis and homology comparison, seven proteins were selected for gene cloning and prokaryotic expression, and the most suitable antigen, TgGRA54, was selected to establish an indirect ELISA for T. gondii. Compared with the indirect immunofluorescent antibody test (IFAT), the positive coincidence rate of the ELISA based on rTgGRA54 was 100% (72/72) and the negative coincidence rate was 80.95% (17/21). The indirect ELISA method based on TgGRA54 recombinant protein was established to detect T. gondii antibodies in bovine sera, and the recombinant protein reacted well with T. gondii positive sera from sheep, mouse, and swine, indicating that the recombinant protein is a good diagnostic antigen for T. gondii.

Anti-Leishmania spp. antibody detection in domestic cats from a visceral leishmaniasis transmission area.

Feline leishmanial infection is reported worldwide, but the epidemiological role of domestic cats in the leishmaniasis cycle remains unclear, and cats might act as cryptic reservoir hosts in endemic areas with no feline leishmaniosis cases. Considering that, a serological screening for anti-Leishmania spp. antibodies was performed by indirect immunofluorescence antibody test (IFAT) in 389 necropsied cats' serum samples from a new visceral leishmaniasis transmission area with no feline leishmanial infection reported to unveil if the cats are being exposed to the parasite. The overall seroprevalence for Leishmania spp. was 11.05% (43/389). No association was found between sex, neutering status, age group, breed, coat length, feline immunodeficiency virus (FIV) infection, and Leishmania spp. antibody detection. A positive association was found with coat color (cats within the orange spectrum with white [particolor]) (OR = 2.47, CI 95% 1 - 6.13, P = 0.044) and a negative association (OR = 0.38, CI 95% 0.18 - 0.79, P = 0.01) between feline leukemia virus (FeLV) infection and IFAT positivity for Leishmania spp. Therefore, it is concluded that the seroprevalence found was greater than 10%, indicating contact of the protozoan with cats in the region served.

One Health Approach in Serosurvey of Toxoplasma gondii in Former Black Slave (Quilombola) Communities in Southern Brazil and Among Their Dogs.

Brazilian quilombos are rural semi-isolated remnant communities of former black slaves and their descendants who traditionally maintained themselves through archaic subsistence livestock and agriculture practices and historically lacked specific public health policies. Although such individuals and their dogs may be exposed to zoonotic pathogens such as Toxoplasma gondii, no study to date has assessed these human-animal populations together. Populations in four different Brazilian quilombos in southern Brazil were evaluated. Overall, 93/208 people (44.7%) and 63/100 dogs (63.0%) were seropositive for IgG anti-T. gondii antibodies by indirect immunofluorescent antibody test (IFAT), 4/208 (1.9%) human samples seropositive for IgM anti-T. gondii antibodies, with a human-dog seropositivity ratio for IgG of 0.71. Quilombola individuals ingesting game meat were 2.43-fold more likely (95% CI: 1.05-5.9) to be seropositive. No risk factors were associated with seropositivity among dogs, thus suggesting that their exposure to T. gondii was random. Surprisingly, our research group had previously found an inverted human-dog ratio for T. gondii seropositivity of 2.54 in the urban area of a nearby major city. Because consumption of raw/undercooked game meat by quilombola individuals may have contributed to higher exposure, higher overall seroprevalence among dogs may have also indicated interaction with wildlife. Although these dogs may hunt wildlife without their owners' awareness, the higher dog seropositivity may also be related to feeding from discarded food in the community or backyard livestock animals and drinking surface water contaminated with oocysts. Thus, wildlife cannot be singled out as the reason, and future studies should consider sampling water, soil, wildlife, and livestock tissues, to fully establish the source of infection in dogs herein.

Choosing Wisely in Immunology Laboratory: Reviewing of Antinuclear Antibody (ANA) Test Requests and Results Together with Other Tests

In this study, it was aimed to examine the distribution and positivity rates of the tests reported in the medical immunology laboratory of a tertiary education and research hospital, and to determine the distribution of antinuclear antibody (ANA) test requests, which constitute a significant part of the workload, according to disease diagnosis/preliminary diagnoses and clinical departments, and to examine the possible causes and costs of incorrect-inappropriate test orders, and to consider effective and applicable solution suggestions. In the study, a retrospective review of data on all immunoserology tests (n=94,954 individual parameters) reported approximately over a three-year period starting from September 2016 was presented. When the tests are divided into three main groups; among all test parameters, ANA tests ranked first with a rate of 20.3% (n=19,248). In our study, the positivity rate of ANA tests evaluated by the indirect immunofluorescence antibody (IFA) method was found as 23.1% (n=4,446). The number of reported parameters of specific autoantibody tests (second group), which include many different tests (IFA, ELISA, and immunoblot based), was 67,976, and the overall positivity was 2.96%, varies between 0.8% and 12.7%, depending on the antibody type. In the ELISA-based brucella and antiviral (herpes simplex virus 1 and 2, varicella virus, measles virus, mumps virus, parvovirus B19) IgM and IgG antibody tests in the third group, the highest positivity rate was observed according to the number of tests requested (30.1%, 2,324/7,730). In ANA-positive patients, the most frequently detected ANA-related autoantibodies were anti-dsDNA (9.2%) and anti-SS-A (8.2%). In ANA-negative patients, in simultaneously ordered tests, anti-dsDNA positivity was found to be 3.3%, while positivity rates for other ANA-related specific autoantibodies were in the range of 0.0-0.31%. ANA tests were most frequently ordered from the rheumatology department (34.2%), and also the highest ANA positivity rate (28%) was observed in this unit. The most notable reason for unnecessary test ordering was the test requests by different physicians for the same patient. We consider that evaluation of ANA tests at a dilution of 1:100 may lead to unnecessary second-step testing (specific autoantibodies) due to the low positive results, and that narrow-scope second-step autoantibody test panels will have negative effects on laboratory efficiency.

Antinuclear antibody staining patterns by indirect immunofluorescence assay observed in patients from a tertiary health center in Latin America

IntroductionThis study aimed to describe the frequency of antinuclear antibody (ANA) staining patterns by indirect immunofluorescence assay observed in patients from a tertiary health center in Latin America. Methodology and methodsThis retrospective, descriptive, and observational study evaluated data from all patients undergoing antinuclear antibody indirect immunofluorescence assay from a single-tertiary center (University Hospital Fundación Valle del Lili, Cali-Colombia) in 2020. ResultsOne thousand and eight patients met the inclusion criteria. The median patient age was 47 (34–59.2) years, and most were female (769, 75.3%). A positive ANA immunofluorescence assay was observed in approximately two-thirds of patients (664, 65.8%). ANA test results were primarily used to exclude a suspected diagnosis in approximately half of the patients (466, 46.2%). Thirty-seven percent (250/664) of the cohort with ANA-positive titers had a systemic autoimmune rheumatic disease (SARD). The most prevalent SARDs included rheumatoid arthritis (RA) (55, 8.2%) followed by systemic lupus erythematosus (SLE) (37, 5.5%). The vast majority of ANA-positive patients had a reported speckled pattern (anti-cell [AC]-2,4,5; 269; 40.5%) followed by homogenous (AC-1; 266; 40%), nucleolar (AC-8,9,10; 46; 6.9%), and centromere (AC-3; 16; 2.4%). The most frequent patterns observed among SLE patients included homogenous (AC-1) patterns in 17 (45.9%) patients, speckled (AC-2,4,5) nuclear patterns in 11 (29.7%) patients, mixed patterns in 7 (18.9%) patients, and reticular/anti-mitochondrial antibody (AMA, AC-21) cytoplasmic patterns in 2 (5.4%) patients. ConclusionsThis study is the first to describe ANA patterns in a Colombian population. Speckled and homogenous patterns were predominant in patients with SARDs.

Detection of bornavirus-reactive antibodies and BoDV-1 RNA only in encephalitis patients from virus endemic areas: a comparative serological and molecular sensitivity, specificity, predictive value, and disease duration correlation study

PurposeHuman Borna disease virus (BoDV-1) encephalitis is an emerging disease in Germany. This study investigates the spectrum of human BoDV-1 infection, characterizes anti-BoDV-1-antibodies and kinetics, and compares laboratory test performances.MethodsThree hundred four encephalitis cases, 308 nation-wide neuropsychiatric conditions, 127 well-defined psychiatric cases from Borna disease-endemic areas, and 20 persons with contact to BoDV-1 encephalitis patients or animals were tested for BoDV-1 infections by serology and PCR.ResultsBoDV-1 infections were only found in encephalitis patients with residence in, or recent travel to, virus-endemic areas. Antibodies were detected as early as 12 days after symptom onset. Serum antibody levels correlated with disease duration. Serology was ordered after 50% of the disease duration had elapsed, reflecting low awareness. BoDV-1-antibodies were of IgG1 subclass, and the epitope on BoDV-1 antigens was determined. Specificity of the indirect immunofluorescence antibody test (IFAT) and lineblot (LB) from serum and cerebrospinal fluid (CSF), as well as PCR testing from CSF, was 100%. Sensitivity, depending on first or all samples, reached 75–86% in serum and 92–94% in CSF for the IFAT, and 33–57% in serum and 18–24% in CSF for the LB. Sensitivity for PCR in CSF was 25–67%. Positive predictive values were 100% each, while negative predictive values were 99% (IFAT), 91–97% (LB), and 90% (PCR). ConclusionsThere is no hint that BoDV-1 causes other diseases than encephalitis in humans. Awareness has to be increased in virus-endemic areas. Tests are robust but lack sensitivity. Detection of IgG1 against specific peptides may facilitate diagnosis. Screening of healthy individuals is likely not beneficial.

Toxoplasma gondii exposure in Brazilian indigenous populations, their dogs, environment, and healthcare professionals

Although Toxoplasma gondii exposure has been reported in indigenous populations worldwide, a One Health approach has not been applied to date. This study concurrently assessed T. gondii exposure in indigenous populations, and their dogs, environment, and indigenous or non-indigenous healthcare professionals (HPs). Human and dog serum samples from 9 indigenous communities in Brazil were assessed by indirect immunofluorescence antibody test for anti-T. gondii antibodies. Soil samples (30 per community) were processed with PCR to amplify T. gondii DNA. Associated risk factors and seroprevalence were analyzed using logistic regression models. Human seropositivity and type of water source were assessed by generalized linear mixed model (GLMM) with binomial error distribution, and game meat consumption with chi-squared test. Overall, 225/463 (49%) indigenous persons were seropositive for anti-T. gondii antibodies. Of all the HPs, 67/168 (40%) were positive, and included 54/147 (37%) positive non-indigenous HPs. Indigenous persons more likely to be seropositive compared with non-indigenous HPs (OR: 1.63; 95% CI: 1.11–2.39). A total of 97/253 (38%) dogs were seropositive and highly associated with seropositive owners (p < 0.001). Based on univariate analysis for indigenous individuals, state location of community (p < 0.001), ethnicity (p < 0.001), consumption of game meat (p < 0.001), type of water source (p < 0.001), and educational level (p = 0.026) were associated with seropositivity. Logistic regression showed that indigenous seropositivity was associated with eating game meat (p = 0.002), drinking water from rivers (p < 0.001), and inversely proportional to the educational level. According to univariate analysis for non-indigenous HP, age (p = 0.005), frequency of visits to the indigenous populations (p < 0.001), consumption of water at the indigenous communities (p < 0.001), and ingestion of raw meat (p = 0.023) were associated with T. gondii seropositivity. Logistic regression revealed living outdoors (p = 0.042), habit of hunting (p = 0.008), and drinking river water (p = 0.007) as risk factors associated to seropositivity in dogs. In addition, indigenous communities lacking water treatment had higher seroprevalence for all groups including indigenous persons (GLMM; z = −7.153; p < 0.001), their dogs (GLMM; z = −2.405; p = 0.0162), and all HPs (GLMM; z = −2.420; p = 0.0155). Human seropositivity was associated with that of their dogs (p < 0.001). A single soil sample, out of 270 (0.37%), was positive for T. gondii by PCR. Our results indicate water source is a risk for human and dog toxoplasmosis in indigenous communities; both share similar exposure. Moreover, quality water access was shown to be crucial to prevent toxoplasmosis in both total and non-indigenous HPs who work in these indigenous communities.

An Investigation of the Relationship Between Isolated Anti-dense Fine-Speckled 70 Autoantibodies and Allergen-Specific Immunoglobulin E.

Background Although the mechanisms of the formation of anti-dense fine-speckled 70 (anti-DFS70) antibodies are not fully known, there is evidence in the literature that allergic reactions may play a role in their formation. Immunoglobulin E (IgE)-mediated immunopathological mechanisms are increasingly being elucidated in diseases such as atopic dermatitis and urticaria-related diseases. We aimed to reveal its relationship with anti-DFS70 in allergen-sensitive patients with positive specific IgE (sIgE) levels. Methodology The study included samples of 758 patients who underwent antinuclear antibody (ANA) screening and allergen-sIgE testing between January 2019 and January 2022. Patients' clinical diagnoses were retrospectively obtained from the hospital information management system. ANA was tested according to the instructions of the manufacturer by the indirect immunofluorescent antibody method using HEp-2 cell substrates (Euroimmun Luebeck, Germany). Allergen-sIgE was determined by chemiluminescence on the Immulite 2000 XPI system (Siemens Healthcare Diagnostics Products GmbH, Marburg, Germany) according to the instructions of the manufacturer. Results ANA pattern was detected in 74 samples included in the study. ANA-positive patients were divided into DFS70 (+) and DFS70 (-) groups. A statistically significant increase in the DFS70 pattern was observed in patients with a positive allergen-sIgE test (p < 0.0001). Both allergen-sIgE and DFS70 positivity were statistically significant in younger age groups (p < 0.05). The most common diagnosis was urticaria-related conditions in 23 (31%) patients with a positive allergy test. Conclusions Our study shows that the positivity of the DFS70 pattern is increased in allergen-sensitive patients. Therefore, the allergen-sIgE-mediated allergic disease should be considered in patients with isolated anti-DSF70. Studies with related disease groups are needed to determine whether there is a relationship between anti-DFS70 and allergy-related disease in these patients. If an immunopathological mechanism is not found, these false-positive results can be considered clinically insignificant, and unnecessary consultations can be avoided.