First laboratory-confirmed case of scrub typhus in Shijiazhuang City, Hebei Province.

Objective: Defining whether a suspected case was due to scrub typhus through laboratory testing, to understand the prevalence of scrub typhus in Shijiazhuang City, Hebei Province. Methods: An epidemiological investigation was conducted on the suspected case, utilizing Weil-Felix test and indirect immunofluorescence assay (IFA) to detect specific antibodies against O. tsutsugamushi in serum specimens. Additionally, PCR amplification of the 56-kDa and groEL genes was performed, followed by constructing a phylogenetic tree to identify the genotype. Results: The acute phase titer of the Weil-Felix test for the case was 1:160, which increased to 1:320 in the recovery phase. IFA assay revealed IgG titers against O. tsutsugamushi of 1:64 in the acute phase and 1:256 in the recovery phase. Sequence alignment of the PCR amplified fragment showed the highest similarity with the O. tsutsugamushi genotype. Kawasaki sequence, ranging from 99.71 to 100.00%. The strain exhibited the closest genetic relationship with the known O. tsutsugamushi Kawasaki genotype. Conclusion: This study confirms the presence of O. tsutsugamushi in Shijiazhuang City, Hebei Province, with the identified strain belonging to the Kawasaki genotype, marking the first diagnosis of this strain in the region.

Clinical Significance of Antibodies to DFS70 in Immunoinflammatory Rheumatic Diseases.

The relevance of the problem of immunoinflammatory rheumatic diseases (IIRD) for modern medicine is determined by their high prevalence in the population, the difficulty of early diagnosis, the rapid development of disability and poor life prognosis. Recent data on the significance of anti-DFS70 have opened up new possibilities for optimizing the step-by-step diagnosis of IIRD. The detection of these antibodies can help in the interpretation of a positive result for antinuclear antibodies (ANA) by indirect immunofluorescence assay on HEp-2 cells (IIFA-HEp-2) in the absence of autoantibodies specific for IIRD. Detection of anti-DFS70 in antinuclear factor (ANF) seropositive patients without clinical and/or serological markers characteristic of a certain disease from the IIRD group can be considered as a potential marker that excludes this group of diseases.

Seasonal Testing, Results, and Effect of the Pandemic on Coxsackievirus Serum Studies.

Coxsackieviruses (CVs) are common causes of infections and can be life-threatening. Unfortunately, rigorous studies guiding the clinician in interpreting CV serum antibody titer testing is lacking. To explore the epidemiology of circulating CVs and the serological test utility in aiding diagnosis of CV infections in our community, we obtained results of CV immunologic diagnostic tests between 2018 and 2022 from a regional healthcare database. For CV type A, rare individuals had positive CF (complement fixation) tests whereas all 16 individuals with IFA testing showed at least one positive serotype. For CV type B CF testing, 52.2% of 222 patients had at least one serotype positive, with B5 being most common and also the most common with higher titers (14.8% with ≥1:32). We found a significant reduction in seropositivity rate during the pandemic in 2020 compared to 2018, which continued through 2022 (OR: 0.2, 95% CI: 0.08-0.49, p-value < 0.001). During the pandemic, the seasonal pattern of positive tests varied from the pre-pandemic pattern. Testing for CVs was increased after the first year of the pandemic. Overall, the variability by month and seasonal change in our data support that CF testing can be used to identify recent CVB infection.

Development of monoclonal antibodies targeting the conserved fragment of hexon protein to detect different serotypes of human adenovirus.

Human adenovirus (HAdV) infects the respiratory system, thus posing a threat to health. However, immunodiagnostic reagents for human adenovirus are limited. This study aimed to develop efficient diagnostic reagents based on monoclonal antibodies for diagnosing various human adenovirus infections. Evolutionary and homology analyses of various human adenoviral antigen genes revealed highly conserved antigenic fragments. The prokaryotic expression system was applied to recombinant penton, hexon, and IVa2 conserved fragments of adenovirus, which were injected into BALB/c mice to prepare human adenovirus-specific monoclonal antibodies. Enzyme-linked immunosorbent assay (ELISA), indirect immunofluorescence assay (IFA), and Western blotting were used to determine the immune specificity of the monoclonal antibodies. Indirect ELISA showed that monoclonal antibodies 1F10, 8D3, 4A1, and 9B2 were specifically bound to HAdV-3 and HAdV-55 and revealed high sensitivity and low detection limits for various human adenoviruses. Western blotting showed that 1F10 and 8D3 specifically recognized various human adenovirus types, including HAdV-1, HAdV-2, HAdV-3, HAdV-4, HAdV-5, HAdV-7, HAdV-21, and HAdV-55, and 4A1 specifically recognized HAdV-1, HAdV-2, HAdV-3, HAdV-5, HAdV-7, HAdV-21, and HAdV-55. IFAs showed that 1F10, 8D3, and 4A1 exhibited highly selective localization to A549 cells infected with HAdV-3 and HAdV-55. Finally, two antibody pairs that could detect hexon antigens HAdV-3 and HAdV-55 at low concentrations were developed. The monoclonal antibodies developed in this study show potential for detecting human adenoviruses. IMPORTANCE: In this study, we selected the three most conserved antigenic fragments of human adenovirus to prepare a murine monoclonal antibody for the first time, and human adenovirus antigenic fragments with heretofore unheard of degrees of conservatism were isolated. The three monoclonal antibodies with the ability to recognize human respiratory adenovirus over a broad spectrum were screened by hybridoma and monoclonal antibody preparation. Human adenovirus infections are serious; however, therapeutic drugs and diagnostic reagents are scarce. Thus, to reduce the serious consequences of human viral infections and adenovirus pneumonitis, early diagnosis of infection is required. The present study provides three monoclonal antibodies capable of recognizing a wide range of human adenoviruses, thereby offering guidance for subsequent research and development.

Serosurvey of Coxiella burnetii in Descendants of Former Black Slaves (Quilombola Communities) of Southern Brazil.

Brazilian descendants of former Black-slave (quilombola) communities have been predisposed to several zoonotic diseases due to social vulnerability, characterized by subsistence and close contact with livestock and companion animals. Accordingly, the present study has assessed anti-Coxiella burnetii antibodies in 200 individuals and 20 dogs from four quilombola communities located in Paraná State, southern Brazil. Serum samples were tested by indirect immunofluorescence assay (IFA) using in-house and commercial diagnostic protocols, with analysis of seropositive titers and antibody type. Fisher's exact test was used to compare seropositivity to C. burnetti with binary variables, with variables with three or more possible responses submitted to logistic regression. In total, 44/200 (22%; 95% CI 16.82-28.24) people tested positive, and 4.5% had titers higher than 128, indicating a recent onset of C. burnetii infection. Seropositive individuals were statistically associated with the Limitão community (p = 0.0013), urban workers as occupations (p = 0.0475), consumption of undercooked meat (p = 0.0159), and contact with animal abortion (p = 0.0276). No seropositivity association was found for age, sex, education, habit of entering forest areas, consumption of game meat, consumption of raw milk, flea and tick bites, dog contact, or history of female miscarriage. Only one of 20 dogs was seropositive with a titer of 128, probably related to an acute animal infection. Despite the prevalence here being higher than previous Brazilian reports, including with symptomatic populations, the results were within range for worldwide outbreaks and occupational risk populations. To the reader's knowledge, this is the first human survey of Q fever in southern Brazil and should be considered a warning for C. burnetii in vulnerable populations, particularly Quilombola communities.

Adopting the International Consensus on ANA Patterns (ICAP) classification for reporting: the experience of Italian clinical laboratories.

The indirect immunofluorescence assay on HEp-2 cells (HEp-2 IFA) is still considered the reference method to detect anti-nuclear antibodies (ANA) because of its high sensitivity and represents a relevant tool for the diagnosis of autoimmune rheumatic diseases. During the last decade, the International Consensus on ANA Patterns (ICAP) initiative promoted harmonization and understanding of HEp-2 IFA staining pattern nomenclature, as well as promoting their use in patient care by providing interpretation for HEp-2 IFA test results. In conjunction with a nationwide survey on the evolution of autoantibody diagnostics in autoimmune rheumatic diseases, we focused on the adherence of the Italian laboratories to the ICAP nomenclature analyzing its lights and shadows. The recent ICAP-oriented report, largely used today among Italian laboratories, also represents a further step in harmonizing and improving communication with the clinicians, adding value to laboratory findings and helping with critical clinical decisions.

Extended autoantibody panel in Turkish patients with early-stage systemic sclerosis: Coexpressions and their influences on clinical phenotypes.

BACKGROUND/AIM: To investigate the frequency and clinical relevance of an extended autoantibody profile in patients with systemic sclerosis (SSc). MATERIALS AND METHODS: In this cross-sectional study, serum from 100 consecutive patients was subjected to indirect immunofluorescence (IIF) (HEp-20-10/primate liver mosaic) and Systemic Sclerosis Profile by EUROIMMUN to evaluate anti-nuclear antibodies (ANA) and autoantibodies against 13 different autoantibodies in patients with SSc less than 3 years. RESULTS: Ninety-three of 100 patients were positive for ANA by IIF. Fifty-three patients showed single positivity, 26 anti-topoisomerase antibodies (anti-Scl70 ab), 16 anticentromere antibodies (ACAs), six anti-RNA polymerase III antibodies (anti-RNAPIII ab), one anti-Ku antibody, one anti-PM/Scl100 antibody, two anti-PM/Scl75 antibodies, one anti-Ro52 antibody, whereas 32 patients had multiple autoantibody positivities. Among classic SSc-specific autoantibodies, anti-Scl70 and anti-RNAPIII abs showed the highest cooccurrence (n = 4). One patient was simultaneously positive for anti-RNAPIII ab and ACA, and one was positive for ACA and anti-Scl70 ab. The clinical features were not statistically different between single and multiple autoantibody-positivity for classic SSc-specific autoantibodies (ACA, anti-Scl70 ab, and anti-RNAPIII ab), except for digital ulcer in the multiantibody positive ACA group (p = .019). CONCLUSION: Based on our results, coexpression of autoantibodies is not uncommon in SSc patients. Although autoantibodies specific to SSc in early disease show generally known clinical features, it remains to be investigated how the coexpression of autoantibodies will affect clinical presentation.

Seroprevalence of Orientia tsutsugamushi and Rickettsia typhi in water buffaloes (Bubalus bubalis) from Southern Thailand.

Background and Aim: Scrub typhus and murine typhus are globally distributed zoonoses caused by the intracellular Gram-negative bacteria Orientia tsutsugamushi and Rickettsia typhi, respectively. Numerous studies have been undertaken on rickettsial illnesses in humans and animals, including arthropod vectors, in Thailand. However, the reports on the seroprevalence of antibodies to O. tsutsugamushi and R. typhi in buffaloes is extremely rare. Thus, this study aimed to estimate the seroprevalence of both rickettsial infections in water buffaloes (Bubalus bubalis) in Phatthalung Province, southern Thailand. Materials and Methods: From February to March 2023, a total of 156 serum samples were collected from 156 water buffaloes on 29 farms in Phatthalung province. The sera were screened for antibodies against O. tsutsugamushi and R. typhi using an indirect immunofluorescence assay. Results: The seroprevalence of antibodies against O. tsutsugamushi and R. typhi in individual water buffaloes was 4.49% (95% confidence interval [CI]: 2.19%-8.97%) and 3.85% (95% CI: 1.77%-8.14%), respectively, whereas 31% (9/29) of the herds had buffaloes with antibodies. The number of buffaloes with scrub typhus infection and ectoparasite infestation was statistically significant (p < 0.05; odds ratio = 6.25 [95% CI: 1.19-33.33]). Intriguingly, the prevalence of scrub typhus antibodies in buffaloes that were not infested with ectoparasites was much higher than those that were. Conclusion: This is the first report of O. tsutsugamushi and R. typhi antibodies in water buffalo sera in Southern Thailand. Two serum samples showed a high antibody titer against O. tsutsugamushi. Seroprevalence mainly occurred in non-ectoparasite-infested buffaloes, especially for O. tsutsugamushi antibodies. At the herd level, one-third of the studied farms showed seroprevalence. Additional research on the occurrence of these pathogens in vectors and in other animal reservoirs is necessary.

Determination of a cut-off value for the serological diagnosis of scrub typhus by detecting anti-Orientia tsutsugamushi immunoglobulin M.

Background & objectives: The diagnosis of scrub typhus (ST) is usually done using enzyme-linked immunosorbent assay (ELISA) due to its ease of performance and reading objectivity. The cut-off value for ELISA needs to be calculated for each geographical location as it depends on zonal endemicity of the disease. This study was, therefore, undertaken to calculate the pan-India cut-off for anti-Orientia tsutsugamushi (OT) immunoglobulin M (IgM) by ELISA. Methods: Samples from cases (cases of ST) and controls (voluntary, consenting, healthy adults) were collected by a network of 29 laboratories across India and tested for anti-OT IgM by immunofluorescence assay (IFA), the considered gold standard test. These samples were retested by ELISA for anti-OT IgM and their optical densities (ODs) were used for cut-off estimation by receiver operating characteristic (ROC) curve. Results: Anti-OT IgM ELISA ODs from 273 controls and 136 cases were used for the cut-off estimation. The ODs of the anti-OT IgM ELISA on healthy individuals and those of confirmed ST cases ranged from 0.1 to 0.75 and 0.5 to 4.718, respectively. ROC curve-based cut-off for ELISA was calculated as 0.554 at a sensitivity of 95.2 per cent and specificity of 95.1 per cent. A value of >1 was noted to have a specificity of 100 per cent in diagnosing ST. Interpretation & conclusions: The cut-off calculated for India was similar to the previous cut-off that was used until now.

Investigation of the Inhibition Activity of Aluminium Oxide Nanoparticles for Herpes Simplex Type 1.

Several studies have shown that Herpes simplex type 1 )HSV-1 (is one of the viruses resistant to medications, so potential antiherpetic agents need to be evaluated. This study aimed to evaluate the impact of Aluminum Oxide Nanoparticles (Al2O3-NPs) on HSV-1 infection. Characterization of Al2O3-NPs was performed using field emission scanning electron microscopy (FESEM), X-ray diffraction (XRD), dynamic light scattering (DLS), and high-resolution transmission electron microscopy (HRTEM). The MTT test was used to investigate the toxicity action of Al2O3-NPs on viable cells. Quantitative Real-Time PCR (qRT-PCR)and TCID50 assays were used to achieve the antiherpetic performance Al2O3-NPs.Indirect immunofluorescence assay (IFA) was performed to determine the inhibitory impact of Al2O3-NPs on viral antigen expression, and acyclovir was utilized as a standard agent in all tests. HSV-1 subjected to Al2O3-NPs at the maximum non-toxic concentration (100 µg / mL) leads to a decrease of 0.1, 0.7, 1.8, and 2.5 log10 TCID50 in the infectious titer relative to virus control (P<0.0001). This concentration of Al2O3-NPs was correlated with 16.9%, 47.1 %, 61.2 %, 72.5 % and 74.6 % inhibition rates, calculated based on HSV-1 viral load compared to virus control. Our results have shown that Al2O3-NPs have a robust antiviral activity against HSV-1. This function demonstrates excellent potential for using Al2O3-NP in topical formulations for treating orolabial or genital herpetic lesions.

Antinuclear antibodies in children: indirect immunofluorescence patterns, antigen targets and associated diagnoses / Anticorps antinucléaires chez l'enfant : aspects en immunofluorescence indirecte, cibles antigéniques et situations cliniques associées

Antinuclear antibodies tests are of a paramount importance in the diagnosis, classification, prognostic evaluation and management of autoimmune diseases in children. The present study aimed to describe the immuno-clinical profile of antinuclear antibodies tests in a pediatric population in order to guide the clinical practice of biologists and clinicians. Our study enrolled 268 children. Antinuclear antibodies screening was performed using the indirect immunofluorescence assay on HEp-2 cells. Identification of target antigens was conducted using separately or at one timepoint the following techniques: enzyme-linked immunosorbent assay, Immunodot and Chemiluminescence. The average age of patients was 9.6 ± 4.3 years, with a female predominance (sex-ratio = 1.9). Antinuclear antibodies screening was positive in 40.67% of cases. The most frequently observed antinuclear antibodies patterns were speckled (52.3%), homogeneous (13.8%) and mixed homogeneous-speckled (13.8%). Autoantibodies were detected in 4 patients (2.51%) for whom ANA testing using the indirect immunofluorescence assay was negative. Positive antinuclear antibodies specificities were detected in connective tissue diseases (44.03%; n = 48), organ-specific autoimmune diseases (10.09%; n = 11), and in non-autoimmune conditions (inflammatory diseases, infections, hematological diseases, vasculitis and Wilson's disease) (32.08%; n = 35). Our study revealed a high rate of positive antinuclear antibodies tests in the pediatric population, mainly related to autoimmune diseases (54.12%) besides non-autoimmune conditions (32.08%). Therefore, screening and interpretation of antinuclear antibodies testing in children require the consideration of clinical data and a close collaboration between clinicians and biologists.

Human Infections with Borna Disease Virus 1 (BoDV-1) Primarily Lead to Severe Encephalitis: Further Evidence from the Seroepidemiological BoSOT Study in an Endemic Region in Southern Germany.

More than 40 human cases of severe encephalitis caused by Borna disease virus 1 (BoDV-1) have been reported to German health authorities. In an endemic region in southern Germany, we conducted the seroepidemiological BoSOT study ("BoDV-1 after solid-organ transplantation") to assess whether there are undetected oligo- or asymptomatic courses of infection. A total of 216 healthy blood donors and 280 outpatients after solid organ transplantation were screened by a recombinant BoDV-1 ELISA followed by an indirect immunofluorescence assay (iIFA) as confirmatory test. For comparison, 288 serum and 258 cerebrospinal fluid (CSF) samples with a request for tick-borne encephalitis (TBE) diagnostics were analyzed for BoDV-1 infections. ELISA screening reactivity rates ranged from 3.5% to 18.6% depending on the cohort and the used ELISA antigen, but only one sample of a patient from the cohort with requested TBE diagnostics was confirmed to be positive for anti-BoDV-1-IgG by iIFA. In addition, the corresponding CSF sample of this patient with a three-week history of severe neurological disease tested positive for BoDV-1 RNA. Due to the iIFA results, all other results were interpreted as false-reactive in the ELISA screening. By linear serological epitope mapping, cross-reactions with human and bacterial proteins were identified as possible underlying mechanism for the false-reactive ELISA screening results. In conclusion, no oligo- or asymptomatic infections were detected in the studied cohorts. Serological tests based on a single recombinant BoDV-1 antigen should be interpreted with caution, and an iIFA should always be performed in addition.

Comparison of Indirect Immunofluorescence and Enzyme Immunoassay for the Detection of Antinuclear Antibodies.

OBJECTIVE: The detection of autoantibodies directed toward nuclear antigens is one of the main criteria for the diagnosis of systemic lupus erythematosus (SLE), for which the most commonly used techniques are the enzyme immunoassay and immunofluorescence assay (IFA). However, the sensitivity and specificity of these tests vary between different techniques. Thus, in this study, we aimed to determine the superior method for detecting antinuclear antibodies (ANAs) and compare the accuracy of tests ordered by rheumatologists versus non-rheumatologists. MATERIALS AND METHODS: We compared the sensitivity and specificity of the two assays in 149 patients from a non-selected population, who were sent to the immunology laboratory of King Abdulaziz Medical City, Jeddah from 2018 to 2019. RESULTS: The sensitivity and specificity of the indirect IFA were 77.78 % and 58.65%, respectively. The positive and negative predictive values of IFA for SLE were 44.87% and 85.92%, respectively. The sensitivity and specificity of the enzyme-linked immunosorbent assay (ELISA) were 77.78% and 80.77%, respectively. The negative and positive predictive values of ELISA for SLE were 63.64% and 89.36%, respectively. The highest number of false-positive IFA tests was requested by family physicians and the lowest was requested by rheumatologists. CONCLUSION: Our data show that IFA has a higher negative predictive value, while ELISA has a higher positive predictive value. The positive predictive value of the test can be improved by pre-selecting patients by specialist rheumatologists.

Case Report: Metagenomic Next-Generation Sequencing Clinches the Diagnosis of Acute Q Fever and Verified by Indirect Immunofluorescence Assay.

Q fever is a zoonotic infectious disease caused by Coxiella burnetii. The clinical symptoms of acute Q fever are usually atypical, and routine serological tests of C. burnetii are not readily available, making the diagnosis of Q fever a challenge. In this case, we report a male patient who had repeated fevers and was administered empirical anti-infective treatment, but the effect was poor. After conducting relevant laboratory and imagological examinations, the etiology has not yet been confirmed. Subsequently, metagenomic next-generation sequencing (mNGS) identified the sequence reads of C. burnetii from the patient's peripheral blood within 48 h, and then the diagnosis of acute Q fever was established. Moreover, the serological test of indirect immunofluorescence assay (IFA) of the C. burnetii antibody was further performed in the Centers for Disease Control, certifying the result of mNGS. The patient was ultimately treated with doxycycline and recovered well. mNGS is an unbiased and comprehensive method in infrequent or culture-negative pathogen identification. To our knowledge, this is the first case of acute Q fever identified by mNGS and confirmed by IFA in Taizhou, China. A further large-scale prospective clinical cohort study is worth carrying out to compare the diagnostic efficiency of mNGS with traditional serological methods and PCR in acute Q fever.

Clinical Performance of Two Automated Immunoassays, EliA CTD Screen and QUANTA Flash CTD Screen Plus, for Antinuclear Antibody Screening.

BACKGROUND: Recently, two fully automated immunoassays for antinuclear antibody (ANA) screening were introduced: EliA CTD Screen (Thermo Fisher Scientific, Freiburg, Germany) and QUANTA Flash CTD Screen Plus (Inova Diagnostics, San Diego, USA). We evaluated their clinical performance in comparison with the indirect immunofluorescence assay (IIFA) and analyzed samples with discrepant results. METHODS: In total, 406 serum samples (206 from patients undergoing routine checkups and 200 from rheumatology clinic patients) were assayed using EliA, QUANTA Flash, and IIFA. We evaluated assay concordance and agreement and confirmed the presence of anti-extractable nuclear antigen (ENA) antibodies in samples with discrepant automated immunoassay and IIFA results. Additionally, we compared the clinical performance of each assay in diagnosing ANA-associated rheumatic disease (AARD) and adjusted the cut-off values. RESULTS: In rheumatology clinic samples, the concordance and agreement were 91.5% and strong between EliA and QUANTA Flash, 79.0% and weak between EliA and IIFA, and 80.5% and moderate between QUANTA Flash and IIFA, respectively. In automated immunoassay-positive, IIFA-negative samples (N=15), all anti-ENA antibodies detected (6/15) were anti-Sjögren's syndrome antigen A/Ro (Ro60) antibodies. The automated immunoassays and IIFA showed high accuracy for diagnosing AARD, and adjusted cut-off values improved their sensitivities (EliA with 0.56 ratio, 82.9% sensitivity; QUANTA Flash with 9.7 chemiluminescent units, 87.8% sensitivity). CONCLUSIONS: The two automated immunoassays showed reliable performance compared with IIFA and can be efficiently used with the IIFA in clinical immunology laboratories. Clinical cut-off values can be adjusted according to the workflow in each laboratory.

Analysis of the impact of sex and age on the variation in the prevalence of antinuclear autoantibodies in Polish population: a nationwide observational, cross-sectional study.

The detection of antinuclear autoantibody (ANA) is dependent on many factors and varies between the populations. The aim of the study was first to assess the prevalence of ANA in the Polish adult population depending on age, sex and the cutoff threshold used for the results obtained. Second, we estimated the occurrence of individual types of ANA-staining patterns. We tested 1731 patient samples using commercially available IIFA using two cutoff thresholds of 1:100 and 1:160. We found ANA in 260 participants (15.0%), but the percentage of positive results strongly depended on the cutoff level. For a cutoff threshold 1:100, the positive population was 19.5% and for the 1:160 cutoff threshold, it was 11.7%. The most prevalent ANA-staining pattern was AC-2 Dense Fine speckled (50%), followed by AC-21 Reticular/AMA (14.38%) ANA more common in women (72%); 64% of ANA-positive patients were over 50 years of age. ANA prevalence in the Polish population is at a level observed in other highly developed countries and is more prevalent in women and elderly individuals. To reduce the number of positive results released, we suggest that Polish laboratories should set 1:160 as the cutoff threshold.

Seroprevalence of rickettsial infection in northern Cyprus: A study among hunters

@#This study was conducted to investigate rickettsial seropositivity among hunters, a high-risk population for tick-borne diseases in northern Cyprus. Serum samples were collected from 300 hunters from different locations during the 2017-2018 hunting season (November 2017 - February 2018). The samples were analyzed by indirect immunofluorescence assay (IFA) using slides coated with Rickettsia slovaca, a species belonging to the spotted fever group (SFG). During the sample collection, a questionnaire was also applied to evaluate possible risk factors for rickettsial seropositivity. Of the 300 serum samples, six (2.0%) were found to be IgG-positive with a titer of 1:64. While all seropositive individuals were male, the statistical analysis revealed no significant association of gender with rickettsial seropositivity (p=1.000). Other factors including age (p=0.414), residential places of the participants (p=0.347), hunting years (p=0.694) or hunting abroad (p=1.000) did not significantly affect the IgG positivity. Also, no statistical correlation was found between a history of an arthropod (tick, louse, or flea) bite and rickettsial seropositivity (p=1.000). To our knowledge, this is the first study that demonstrates rickettsial seropositivity among human population in northern Cyprus. Our study suggests that awareness should be raised among the people especially involved in outdoor activities such as hunting, and control programs should be implemented to prevent possible rickettsiosis cases. Further serological studies using other Rickettsia spp. antigens, as well as molecular studies that search for Rickettsia spp. in humans, animals and arthropods are needed to obtain more comprehensive data on rickettsiosis in northern Cyprus.

Seroprevalence and hematological abnormalities associated with Ehrlichia canis in dogs referred to a veterinary teaching hospital in central-western Brazil / Soroprevalência e alterações hematológicas associadas com Ehrlichia canis em cães encaminhados a um hospital veterinário-escola no centro-oeste do Brasil

Ticks are significant parasites of dogs in the tropics, where tick-borne pathogens are highly prevalent, especially in areas where tick control measures are frequently neglected. This study investigated the seroprevalence and hematological abnormalities associated with Ehrlichia canis in dogs referred to a veterinary teaching hospital in Central-western Brazil. Out of 264 dogs tested for anti-Ehrlichia canis antibodies by an indirect immunofluorescence assay (IFA), 59.1% (156/264) were positive. Seropositivity was significantly associated to anemia and thrombocytopenia, alone or in combination, and to leukopenia. Conversely, there were no differences in terms of seroprevalence according to sex, breed and age. This study demonstrated that dogs referred to a veterinary teaching hospital in Central-western Brazil are highly exposed to E. canis and that seropositive dogs are more likely to present hematological abnormalities, particularly anemia, thrombocytopenia and leukopenia. To our knowledge, this is the first study on detection of anti-E. canis antibodies by means of IFA among dogs in the state of Goiás. These findings highlighted the need for increasing awareness among dog owners regarding tick control measures in Central-western Brazil, ultimately to reduce the risk of exposure to E. canis and other tick-borne pathogens.

Carrapatos são importantes parasitos de cães nos trópicos, onde patógenos transmitidos por carrapatos são altamente prevalentes, especialmente em áreas onde as medidas de controle de carrapatos são frequentemente negligenciadas. O estudo investigou a soroprevalência e as anormalidades hematológicas associadas à Ehrlichia canis em cães encaminhados para um hospital veterinário-escola no Centro-oeste do Brasil. Dos 264 cães testados para anticorpos anti-Ehrlichia canis por meio da reação de imunofluorescência indireta (RIFI), 59.1% (156/264) foram positivos. A soropositividade foi associada significativamente à anemia e trombocitopenia, isoladamente ou em combinação, e à leucopenia. Por outro lado, não houve diferenças quanto à soroprevalência segundo sexo, raça e idade. Este estudo demonstrou que cães encaminhados a um hospital veterinário-escola na região Centro-oeste do Brasil são altamente expostos à E. canis, e que cães soropositivos têm maior probabilidade de apresentar alterações hematológicas, principalmente anemia, trombocitopenia e leucopenia. Para o nosso conhecimento, este é o primeiro estudo sobre a detecção de anticorpos anti-E. canis por meio da RIFI em cães do estado de Goiás. Essas descobertas destacam a necessidade de aumentar a conscientização entre os proprietários de cães em relação às medidas de controle do carrapato no Centro-oeste do Brasil, em última análise, para reduzir o risco de exposição ao E. canis e outros patógenos transmitidos por carrapatos.

Preparation and Characterization of a New Monoclonal Antibody Specific Against Lawsonia intracellularis and Its Application in Indirect Immunofluorescence and Immunocytochemistry Assay.

Proliferative enteropathy (PE) is an infectious enteric disease caused by Lawsonia intracellularis (L. intracellularis) and is endemic in pig herds worldwide. However, a L. intracellularis-specific monoclonal antibody plays an important role in the evaluation of L. intracellularis infection in vitro. Therefore, the objective of this study was to produce and identify the characteristics of a new monoclonal antibody against the outer membrane protein (Omp2) of L. intracellularis and apply it in an indirect immunofluorescence assay (IFA) and immunocytochemistry (IHC). The results indicated that three highly specific monoclonal antibodies against the Omp2 protein (4D9, 3G2, and 7G5) of L. intracellularis were obtained by using purified Omp2 as an immunogen, the titers of ascitic fluids of 4D9, 3G2, and 7G5 cells were 1:2,048,000, 1:512,000, and 1:256,000, respectively. IFA analysis showed that the 4D9, 3G2, and 7G5 have no cross-reactivity with other enteric bacteria commonly found in the ilea of pigs or closely related to L. intracellularis, such as Desulfovibrio, Bilophila wadsworthia (B. wadsworthia), Salmonella choleraesuis (S. choleraesuis), Salmonella typhimurium (S. typhimurium), Escherichia coli (E. coli), and Brachyspira hyodysenteriae (B. hyodysenteriae). IFA and IHC results indicated that the monoclonal antibodies can be successfully used as primary antibodies to detect L. intracellularis in infected cells and in the crypt of the ileum from infected tissues of PE. Our findings suggested that the new monoclonal antibody specific against L. intracellularis will be useful for the evaluation of L. intracellularis infection in vivo and in vitro.