Occurrence of Coxiellosis in ruminants and its associated risk factors.

Coxiellosis in animals is caused by the zoonotic pathogen, Coxiella burnetii. Although the disease is of public health importance it remains underdiagnosed and underreported. The cross- sectional study was aimed to estimate the occurrence of the disease in livestock of study area and also to identify the risk factors associated with the disease in animals. Blood, serum, and vaginal swabs samples were collected from 200 ruminants (cattle, sheep, and goats), across various farms in Karnataka, India. These samples were then screened using ELISA and PCR (com1 and IS1111). A questionnaire was administered to the farm owners to collect the risk factor-related information. About 5.26 % cattle, 12.3 % sheep, and 12.5 % goats were positive by ELISA. By PCR, 9.47 % cattle, 9.3 % sheep, and 10 % goats were positive. Overall, the occurrence of 14.73 %, 18.46 % and 17.5 % was estimated in cattle, sheep and goat, respectively. PCR targeting the IS1111 gene detected higher number of samples as positive as compared to the com1 gene PCR. Higher number of vaginal swab samples were detected as positive as compared to blood. History of reproductive disorders (OR: 4.30; 95 %CI:1.95- 9.46), abortion (OR: 30.94; 95 %CI:6.30- 151.84) and repeat breeding (OR:11.36; 95 %CI:4.16- 30.99) were significantly associated with coxiellosis (p < 0.005). Multivariable analysis by logistic regression model analysis suggested retained abortion, repeat breeding and rearing of animal in semi-intensive system as factors significantly associated with the infection. Cultural identification of the PCR positive samples were cultured using embryonated egg propagation and cell culture techniques and positivity was confirmed in six samples. Phylogenetic analysis of the com1 and IS1111 gene revealed clustering based on similar geographic locations. The study estimated the occurrence of the disease in the study area and identified the potential risk factors.

Comparative analysis of diagnostic assays for scrub typhus: Unveiling enhanced approaches for accurate detection.

The study comparatively evaluated serological assays, namely, Weil Felix assay, and IgM ELISA with the gold-standard immunofluorescence test (IFAT) for the sensitive and specific serodiagnosis of scrub typhus infection in occupationally exposed groups of humans. A total of 78 serum samples collected from persons affected with various ailments and belonging to different risk groups were screened in the study. Out of the 78 serum samples tested, a total of 17, 26, and 47 samples turned out to be positive by IFAT, IgM ELISA, and Weil Felix test, respectively. The Weil Felix assay could not serve as an ideal test for screening scrub typhus infection owing to its poor sensitivity and specificity in comparison with IFAT. IgM-ELISA could be an initial screening test to detect scrub typhus suspected patient in limited resource settings.

Serological and molecular detection of Toxoplasma gondii and Neospora caninum in free-ranging rats from Nagpur, India.

Toxoplasma gondii and Neospora caninum are cyst-forming coccidian parasites that infect both wild and domestic non-felids as intermediate hosts, with rodents serving as important reservoir hosts during their life cycles. This study was aimed at investigating T. gondii and N. caninum infections and identifying factors favouring T. gondii infection in free-ranging rats from India. A total of 181 rodents were trap-captured, and blood and brain samples were subsequently collected for serological and molecular examination of T. gondii and N. caninum. Antibodies against T. gondii and N. caninum were detected by MAT/NAT and IFAT in 13.8% (25/181) and 1.65% (3/181) of rodents, respectively. All three N. caninum samples positive by NAT/IFAT were also positive for ELISA, while for T. gondii, 19 of 25 MAT/IFAT positive samples were also positive for ELISA. The antibody titers (MAT/NAT/IFAT) of rodents seropositive for T. gondii ranged from 25 to 400, while those of rats seropositive for N. caninum ranged from 25 to 100. Also, using PCR, DNA from T. gondii (B1 gene) and N. caninum (NC5 gene) was found in 2.76% (5/181) of brain samples and 0.55% (1/181) of brain samples. All PCR positive samples were also seropositive. No mixed infections were observed in the serological and molecular detections. A Chi-square analysis revealed that older rats and rats living in urban areas are significantly associated with T. gondii infection; however, rodent species, gender, location, habitat types, and seasonality were statistically nonsignificant. Overall, this study demonstrated that T. gondii was widely distributed while N. caninum was less prevalent among free-ranging rats in the studied area.

Edifications on Indirect IgM ELISA and Immunofluorescence assay (IFA) of scrub typhus in humans and erudition of Orientia tsutsugamushi in the vector of cohabiting rodents.

Scrub typhus is a vector-borne disease caused by Orientia tsutsugamushi, propagated into humans by the bite of infected mite belonging to genus Leptotrombodium. The present study was conducted in the Nagpur region of central India aiming towards a survey of cohabiting rodents and their potential vectors for the presence of Orientia tsutsugamushi by PCR method. The study also emphasizes serological diagnosis of the disease by employing indirect IgM ELISA and IFA amongst the human cases of pyrexia of unknown origin. Indirect IgM ELISA recorded 39.69% (31/92) seropositive patients, further processing of ELISA positive samples for IFA revealed 67.74 % (21/31) positivity for Boryong, Gilliam, Karp, and Kato serotypes. A total of 50 rodents were trapped from the cohabit areas of the patients. Three different types of rodents were identified; among which, Rattus bandicoot was highest. From these rodents, 164 vectors viz mites, lice, and fleas were collected. The highest chiggar index was recorded for Ornithonyssus biscotti mites (3.4). This study prompts a detailed analysis of different species of rodents and vectors in the said endemic region.

Optimization of In-House Indirect-ELISA & EITB Assays Employing Cysticercus cellulosae Antigens for Serological Detection and PCR Assays for Molecular Detection of Porcine Cysticercosis.

Background: Porcine cysticercosis, caused by metacestodes of Taenia solium is an important neglected zoonosis. We evaluated the presence of anti-cysticercal antibodies and T. solium specific DNA in pig sera and blood samples respectively collected from Maharashtra, India. Methods: A total of three antigens (Scolex Antigen (SA), Membrane Body Antigen (MBA) and Excretory-Secretory Antigen (ESA)) were prepared from metacestodes of T. solium and employed in an in-house developed indirect-IgG ELISA for serological screening of 1000 porcine sera samples at Department of Veterinary Public Health, Nagpur Veterinary College, Maharashtra, India. The ELISA positive sera samples were subjected to EITB Assay for detection of immunodominant peptides. An effort has been made for molecular detection of porcine cysticercosis by PCR assay targeting large subunit rRNA gene of T. solium from blood samples of the corresponding ELISA-positive pigs. Results: The overall seroprevalence of porcine cysticercosis employing SA, MBA and ESA was 12.6%, 8.7% and 12.5% respectively. The lower and medium molecular weight peptides were the most frequently recognised in EITB assay. The numbers of bands recognised in EITB assay were observed to be proportionate with the corresponding ELISA O.D. values. An amplification product of 286 bp was observed in 22.98% (20/87), 30.35% (30/99) and 17.14% (12/70) of the sero-positive samples against SA, ESA and MBA respectively. Conclusion: EITB still remains the gold standard serodiagnosis test for cysticercosis. The inclusion of a greater number of positive samples and purification of antigens may improve the diagnostic efficacy of the tests.

Seroprevalence, risk factors, and serological cross-reactivity for diagnosis of Toxoplasma gondii and Neospora caninum infections in goats in India.

Toxoplasma gondii and Neospora caninum are genetically related cyst-forming protozoan parasites that cause reproductive failures in ruminants. Given the limited information on the epidemiology of these infections in goats in India, the study aimed to estimate the seroprevalence, assess antibody cross-reactivity for diagnosis, and identify associated risk factors. A total of 695 sera were evaluated for antibodies to T. gondii and N. caninum infections using Modified Agglutination Test (MAT for Toxoplasma)/Neospora agglutination test (NAT), Enzyme-linked immunosorbent assay (ELISA), and Indirect Fluorescent Antibody Test (IFAT for tachyzoite and bradyzoite stages). The seroprevalence rate of T. gondii and N. caninum infections was 56.9% and 10.9%, respectively. Inter-rater agreement (kappa value - κ) was calculated to assess agreements between various diagnostic assays, using the IFAT as the gold standard (for detecting both infections), the agreements for MAT/NAT (κ = 0.97) and the ELISA (κ = 0.95) were excellent. The acute infection among seropositive goats were determined using serological (IgG avidity test - measures the binding strength between IgG antibodies and parasite antigens) and molecular diagnoses (PCR for repetitive DNA sequences - Toxoplasma B1 gene: 131 bp and Neospora NC5 gene: 328 bp). Among seropositive goats ≥80% had high IgG avidity and <10% of animals had low IgG avidity antibodies for both infections. Most low IgG avidity goats were PCR positive for the TgB1 gene (94.4%) or Nc5 gene (85.7%). In the serological assays, we used different dilutions of test serum to rule out the cross-reactivity owing to similar antigenic makeup between these two parasites. When the serological cross-reactivity was analyzed using invasion assay at a serum titer of ≥200, more than 90% T. gondii positive sera showed host cell invasion of N. caninum and vice versa. Largely, the serological results indicate that cut-off serum dilution of ≥1:200 for ELISA and IFAT and ≥1:25 for MAT/NAT avoids serological cross-reactivity between T. gondii and N. caninum. Further, the Univariate and multivariate analyses showed that adult animals (>2 years), reservoir hosts, and extensive rearing systems are common risk factors for these infections. However, the history of abortion was identified as a significant risk factor for T. gondii infection. This study revealed that T. gondii and N. caninum infections are highly prevalent in this region and the use of an appropriate cut-off serum dilution is necessary to avoid cross-reactivity between these closely related parasites.

Seroprevalence and associated risk factors of Toxoplasma gondii and Neospora caninum infections in cattle in Central India.

Toxoplasma gondii and Neospora caninum are closely related cyst-forming parasites identified as important causes of reproductive failures in ruminants. While these parasites have been reported worldwide, seroprevalence and associated risk factors for cattle infections have not been determined in India. A total of 576 serum samples of cattle were analyzed for antibodies to T. gondii and N. caninum using enzyme-linked immunosorbent assay (ELISA), modified/Neospora agglutination test (MAT/NAT), and an indirect fluorescent antibody test (IFAT-tachyzoite and bradyzoite). Additionally, general information about cattle, movement of cats and dogs, the menace of rodents, management, and reproductive disorders were assessed to identify the potential risk factors. Overall, 32.9% (190/576) serum samples reacted positively to T. gondii and 24.8% (143/576) to N. caninum. The performance of the diagnostic tests showed excellent agreement between IFAT and ELISA (kappa [κ] = 0.98) and between MAT/NAT and ELISA (κ = 0.97). Combining both infections on avidity test, 94% sera had high-IgG avidity, and 3% had low-IgG avidity antibodies, indicating chronic infection in the majority of the cases. The identified risk factors (p < 0.05) for exposure to T. gondii were: increasing age (Odds Ratio [OR]: 2.02), movement of cat (OR: 4.8) and rodents (OR: 1.57) in the farm; and for N. caninum: increasing age (OR: 1.6), movement of dogs in the farm (OR: 2.07), drinking pond water (OR: 1.64) and abortion (OR: 1.82). These findings revealed that T. gondii and N. caninum infections are widespread in the study area and suggest conducting nationwide epidemiological studies owing to their economic importance.

Seroprevalence and risk factors of Toxoplasma gondii infection among veterinary personnel and abattoir workers in Central India.

Toxoplasmosis, caused by the protozoan parasite Toxoplasma gondii, is an important zoonotic infection. Veterinary personnel and abattoir workers are considered to be at a high risk of T. gondii infection owing to their occupational exposure. However, the association of T. gondii infection with occupational exposure to animals has not been determined in India. Hence, we analysed 139 and 126 blood samples of veterinary personnel and abattoir workers, respectively, for anti-T. gondii antibodies using enzyme-linked immunosorbent assay (ELISA), modified agglutination test (MAT) and indirect fluorescent antibody test (IFAT). The association of seroprevalence with sociodemographic profiles, work activities and dietary habits was determined in the study population. MAT, ELISA and IFAT results demonstrated nearly 46%, 48% and 47% seropositivity, respectively. MAT (kappa = 0.924) and IFAT (kappa = 0.962) results showed good agreement with ELISA results. Of the ELISA positive samples, 46% was copositive for IgG antibody, 1.5% for IgM antibody and 1.5% for both IgG and IgM antibodies. High IgG avidity was observed only in IgG+ IgM- and IgG+ IgM+ samples and not in IgM+ IgG- samples, indicating chronic T. gondii infection in most of the cases. Furthermore, multivariate analysis revealed that T. gondii seropositivity was associated with age > 30 years (odds ration [OR] = 1.992), cat at home (OR = 1.991), not wearing gloves (OR = 1.886), not wearing safety glasses (OR = 1.985) and contact with soil (OR = 1.695). These findings support the presence of a potentially significant association between T. gondii seropositivity and occupational exposure to animals.

Toxoplasma gondii induces robust humoral immune response against cyst wall antigens in chronically infected animals and humans.

Toxoplasma gondii differentiation from proliferating tachyzoites into latent bradyzoites is central to pathogenesis and transmission. Strong humoral immune response has been reported against tachyzoite antigens, however, antibody-mediated response towards bradyzoite antigens is poorly characterized. This work aimed to study the humoral immune response towards bradyzoite and associated cyst wall antigens particularly CST1. The immunoreactivity of 404 goats, 88 sheep and 92 human sera to recombinant (CST1 and SRS9) and native proteins of encysted bradyzoite along with well-established tachyzoite antigens (SAG1 and GRA7) was determined using ELISA, Western blot and immunofluorescence analysis (IFA). ELISA results revealed nearly 50% of sera contain T. gondii specific antibodies. Results were further validated using Western blot and IFA. T. gondii positive sera predominantly recognized the cyst wall besides the known tachyzoite surface antigens. The presence of CST1 antibodies in seropositive samples were in line with the staining patterns which were consistent with CST localization. Notably, T. gondii IgM- IgG+ sera recognize the cyst wall whereas IgM + IgG-sera recognize tachyzoite antigens indicating acute infection consistent with presence of parasite DNA. The study demonstrates a strong humoral response against bradyzoite associated cyst wall antigens across naturally infected animals and humans. CST1 emerged as a key immunomodulatory antigen which may have direct implications for clinical immunodiagnostics.