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Vesicular stomatitis virus (VSV) outbreaks periodically occur in livestock in the western US and are thought to originate from outside this country. Feral swine (Sus scrofa) have been identified as an amplifying host for vesicular stomatitis New Jersey virus (VSNJV) and have been used to better understand the epidemiology of this virus through serosurveillance. This study aimed to determine if antibodies to vesicular stomatitis Indiana virus (VSIV) and VSNJV were present in feral swine in the western US and to determine if seropositive animals were associated with areas of previously detected VSV in livestock. A total of 4,541 feral swine samples was tested using virus neutralization (VN); samples exhibiting neutralizing activity against one or more of the viruses were confirmed using competitive ELISA (cELISA). Eight sera exhibited neutralizing activity by VN assay and a single serum sample from an animal from Kinney County, Texas sampled in December 2019 tested positive for antibodies to VSIV by cELISA. This finding is supported by a local outbreak of VSIV in horses in the same county in June 2019. The low prevalence of antibodies against VSNJV and VSIV was unexpected but indicates that feral swine in the western US do not represent an endemic reservoir for either of these viruses. [ABSTRACT FROM AUTHOR]

Abstract Contagious caprine pleuropneumonia (CCPP) is a severe and devastating respiratory disease of goats, which is characterized by severe serofibrinous pleuropneumonia accompanied by high morbidity and mortality. A cross-sectional study was conducted from July 2022 to January 2023 to determine the seroprevalence of CCPP and identify risk factors associated with the occurrence of CCPP in goats in five selected districts of the South Wollo Zone of the Eastern Amhara region. A total of 384 sera samples were collected from goats and examined for antibodies specific to Mycoplasma capricolum subspecies capripneumoniae (Mccp) using Competitive Enzyme-Linked ImmunoSorbent Assay (cELISA) test. Out of the total examined sera, 26 samples were positive for CCPP, giving an overall seroprevalence of 6.7% (95% CI = 6.64–9.77). A seroprevalence of 5.05%, 4.65%, 2.78%, 12.90%, and 10.77% were recorded in Ambasel, Tehuledere, Kalu, Dessie Zuria and Kutaber districts, respectively. However, there was no statistically significant difference among these five districts (p > 0.05). The seroprevalence of CCPP varies significantly between age groups and agroecology (p 0.05). Old-aged goats (OR = 4.10) and goats found in the lowlands (OR = 5.09) were at higher risk of infection with CCPP than young-aged goats and goats found in the highlands, respectively. In conclusion, the present seroprevalence investigation indicated the occurrence of CCPP in those selected study districts of the South Wollo Zone. Therefore, appropriate control measures, including avoiding the mixing of flocks and vaccination should be designed and implemented especially in the lowland areas and older goats to reduce the further spread and magnitude of the disease.

Abstract Lumpy skin disease (LSD) is a highly contagious disease caused by lumpy skin disease virus (LSDV) in bovines. Rapid and accurate diagnosis is very important to controll it. However, current commercial detection kits need to be improved in terms of sensitivity or specificity. This study aimed to develop a novel diagnostic competitive enzyme-linked immunosorbent assay (cELISA) based on the newly identified antigen gene LSDV034. The rLSDV034 protein was identified as a potential diagnostic antigen, and it was expressed, purified, and used to immunize BALB/c mice. Using laboratory-prepared indirect ELISA (iELISA), the positive cell lines were screened, and their blocking activity was further verified by competitive ELISA (cELISA). The cell line, 1H7, was chosen to produce mouse ascites, which were purified for a monoclonal antibody (mAb, 5.395 mg/mL). The heavy chain type of the 1H7 mAb was identified as IgG1a, and its light chain subtype was identified as κ. Furthermore, cELISA was developed to detect bovine serum antibodies, with rLSDV034 (4 μg/mL) as the coating antigen and HRP-1H7 mAb (1:300) as the competitive antibody. The cutoff value of cELISA was 55%, based on 32 negative bovine serum samples. The analytical sensitivity was 1:8, and no cross-reaction was detected with bovine viral diarrhea virus (BVDV), infectious bovine rhinotracheitis virus (IBRV), Pasteurella multocida (P. multocida), or Mycoplasma bovis (M. bovis) from the serum samples. The diagnostic sensitivity and specificity of cELISA were 98.46% (95% confidence interval, CI: 91.7–100) and 100% (95% CI: 89.1–100), respectively, based on the analysis of 30 LSDV-infected bovine serum samples, 35 GTPV-vaccinated samples, and 32 negative samples. The overall coincidence of the cELISA with the virus neutralization test (VNT) reached 98.97% (95% CI: 94.4–100). Furthermore, we used cELISA to analyze 230 clinical bovine serum samples (including 59 infected and 171 vaccinated samples) and found that the serum positivity rates of the immunized samples (on d 60 postimmunization) and infected samples were 77.78% (95% CI: 70.8–83.8%) and 71.19% (95% CI: 57.9–82.2), respectively. These results indicate that the developed cELISA is promising for detecting serum antibodies in naturally infected or vaccinated cattle.

Contagious caprine pleuropneumonia (CCPP) is a severe and devastating respiratory disease of goats, which is characterized by severe serofibrinous pleuropneumonia accompanied by high morbidity and mortality. A cross-sectional study was conducted from July 2022 to January 2023 to determine the seroprevalence of CCPP and identify risk factors associated with the occurrence of CCPP in goats in five selected districts of the South Wollo Zone of the Eastern Amhara region. A total of 384 sera samples were collected from goats and examined for antibodies specific to Mycoplasma capricolum subspecies capripneumoniae (Mccp) using Competitive Enzyme-Linked ImmunoSorbent Assay (cELISA) test. Out of the total examined sera, 26 samples were positive for CCPP, giving an overall seroprevalence of 6.7% (95% CI = 6.64–9.77). A seroprevalence of 5.05%, 4.65%, 2.78%, 12.90%, and 10.77% were recorded in Ambasel, Tehuledere, Kalu, Dessie Zuria and Kutaber districts, respectively. However, there was no statistically significant difference among these five districts (p > 0.05). The seroprevalence of CCPP varies significantly between age groups and agroecology (p < 0.05). However, the seroprevalence did not vary with sex, body condition score (BCS), and flock size (p > 0.05). Old-aged goats (OR = 4.10) and goats found in the lowlands (OR = 5.09) were at higher risk of infection with CCPP than young-aged goats and goats found in the highlands, respectively. In conclusion, the present seroprevalence investigation indicated the occurrence of CCPP in those selected study districts of the South Wollo Zone. Therefore, appropriate control measures, including avoiding the mixing of flocks and vaccination should be designed and implemented especially in the lowland areas and older goats to reduce the further spread and magnitude of the disease. [ABSTRACT FROM AUTHOR]

Lumpy skin disease (LSD) is a highly contagious disease caused by lumpy skin disease virus (LSDV) in bovines. Rapid and accurate diagnosis is very important to controll it. However, current commercial detection kits need to be improved in terms of sensitivity or specificity. This study aimed to develop a novel diagnostic competitive enzyme-linked immunosorbent assay (cELISA) based on the newly identified antigen gene LSDV034. The rLSDV034 protein was identified as a potential diagnostic antigen, and it was expressed, purified, and used to immunize BALB/c mice. Using laboratory-prepared indirect ELISA (iELISA), the positive cell lines were screened, and their blocking activity was further verified by competitive ELISA (cELISA). The cell line, 1H7, was chosen to produce mouse ascites, which were purified for a monoclonal antibody (mAb, 5.395 mg/mL). The heavy chain type of the 1H7 mAb was identified as IgG1a, and its light chain subtype was identified as κ. Furthermore, cELISA was developed to detect bovine serum antibodies, with rLSDV034 (4 μg/mL) as the coating antigen and HRP-1H7 mAb (1:300) as the competitive antibody. The cutoff value of cELISA was 55%, based on 32 negative bovine serum samples. The analytical sensitivity was 1:8, and no cross-reaction was detected with bovine viral diarrhea virus (BVDV), infectious bovine rhinotracheitis virus (IBRV), Pasteurella multocida (P. multocida), or Mycoplasma bovis (M. bovis) from the serum samples. The diagnostic sensitivity and specificity of cELISA were 98.46% (95% confidence interval, CI: 91.7–100) and 100% (95% CI: 89.1–100), respectively, based on the analysis of 30 LSDV-infected bovine serum samples, 35 GTPV-vaccinated samples, and 32 negative samples. The overall coincidence of the cELISA with the virus neutralization test (VNT) reached 98.97% (95% CI: 94.4–100). Furthermore, we used cELISA to analyze 230 clinical bovine serum samples (including 59 infected and 171 vaccinated samples) and found that the serum positivity rates of the immunized samples (on d 60 postimmunization) and infected samples were 77.78% (95% CI: 70.8–83.8%) and 71.19% (95% CI: 57.9–82.2), respectively. These results indicate that the developed cELISA is promising for detecting serum antibodies in naturally infected or vaccinated cattle. [ABSTRACT FROM AUTHOR]

In November 2021, an investigation was conducted into an outbreak of abortion, stillbirth, and the birth of calves with congenital abnormalities (arthrogryposis and hydranencephaly) at a dairy farm in Dasht-e-Mughan city, Ardabil province. A total of 70 cows experienced these issues. To determine the cause of the outbreak, post-mortem brain tissue samples were collected from two calves affected by hydranencephaly, which occurred shortly after their birth. Polymerase chain reaction (PCR) testing was conducted for multiple viruses, including bovine viral diarrhea (BVD), border disease, Akabane, Schmallenberg, and bluetongue viruses (BTVs). The samples were positive only for Akabane virus. Serum samples were collected from a group of 60 cattle, consisting of 45 adult cows and 15 younger calves aged between 8 to 10 months. These samples were analyzed to detect the presence of antibodies against the Akabane and Schmallenberg viruses. Both of these viruses are known to be responsible for causing abortion, stillbirth, and congenital abnormalities in calves. Among 45 cows that tested by competitive enzyme-linked immunosorbent assay (cELISA), 26.66% and 33.33% exhibited antibodies against Akabane and Schmallenberg viruses, respectively. Notably, 20.00% of cows co-exhibited antibodies for both viruses. Despite PCR evidence implicating Akabane virus as the principal etiology of clinical signs observed in the affected herd, the high coseropositivity to Schmallenberg virus, warrants a thorough investigation into potential viral interactions. Further research is required to determine the source of the virus and their transmission routes. This information could facilitate the refinement of disease control strategies and improving the management of reproductive challenges in such affected herds. [ABSTRACT FROM AUTHOR]

A cross-sectional study was conducted to determine the seroprevalence of the Peste des Petits Ruminant (PPR) virus (PPRV) in sheep populations and to determine the potential epidemiological risk factors associated with this infection. Between October 2014 and March 2017, 2420 sheep serum samples were collected from ten selected PPR outbreak-prone districts in Bangladesh. The collected sera were analysed by competitive enzyme-linked immunosorbent assay (cELISA) test to detect antibodies against PPR. A previously designed disease report form was used to gather data on important epidemiological risk factors, and a risk analysis was performed to ascertain their association with PPRV infection. By cELISA, 44.3 % (95 % confidence interval:42.4-46.4 %) of sheep sera were positive for PPRV antibodies against PPR. In univariate analysis, the Bagerhat district had significantly higher seropositivity (54.1 %, 156/288) than other districts. Moreover, significantly higher (p

Abstract Background Tick-borne encephalitis (TBE) is a severe human neuroinfection caused by TBE virus (TBEV). TBEV is transmitted by tick bites and by the consumption of unpasteurized dairy products from infected asymptomatic ruminants. In France, several food-borne transmission events have been reported since 2020, raising the question of the level of exposure of domestic ungulates to TBEV. In this study, our objectives were (i) to estimate TBEV seroprevalence and quantify antibodies titres in cattle in the historical endemic area of TBEV in France using the micro virus neutralisation test (MNT) and (ii) to compare the performance of two veterinary cELISA kits with MNT for detecting anti-TBEV antibodies in cattle in various epidemiological contexts. A total of 344 cattle sera from four grid cells of 100 km² in Alsace-Lorraine (endemic region) and 84 from western France, assumed to be TBEV-free, were investigated. Results In Alsace-Lorraine, cattle were exposed to the virus with an overall estimated seroprevalence of 57.6% (95% CI: 52.1–62.8%, n = 344), varying locally from 29.9% (95% CI: 21.0–40.0%) to 92.1% (95% CI: 84.5–96.8%). Seroprevalence did not increase with age, with one- to three-year-old cattle being as highly exposed as older ones, suggesting a short-life duration of antibodies. The proportion of sera with MNT titres lower than 1:40 per grid cell decreased with increased seroprevalence. Both cELISA kits showed high specificity (> 90%) and low sensitivity (less than 78.1%) compared with MNT. Sensitivity was lower for sera with neutralising antibodies titres below 1:40, suggesting that sensitivity of these tests varied with local virus circulation intensity. Conclusions Our results highlight that cattle were highly exposed to TBEV. Screening strategy and serological tests should be carefully chosen according to the purpose of the serological study and with regard to the limitations of each method.

Abstract Background Contagious bovine pleuropneumonia (CBPP) is an economically important infectious disease that is characterized by a variable course and insidious nature. A cross-sectional study was conducted in El Jazeera State, Central Sudan, to determine the seroprevalence and risk factors of CBPP in cattle from seven localities. A total of 218 serum samples were randomly collected from apparently healthy cattle aged older than 6 months between April and May 2021 and were tested serologically using a commercial ELISA kit. Results The overall seroprevalence of CBPP was 50.5% (110/218). Univariate analysis showed a significant difference (p

Abstract Background Contagious bovine pleuropneumonia [CBPP] is a transboundary animal disease of cattle caused by Mycoplasma mycoides subsp. mycoides [Mmm]. CBPP causes severe economic losses to livestock producers in sub-Saharan Africa mainly due to high mortality, morbidity, reduction in productivity as well as livestock trade restrictions. This study aimed at determining seroprevalence of Mmm in cattle from Karamoja region, north-eastern Uganda; data that are required to design and implement risk based CBPP control program. Methods We randomly collected blood samples from 2,300 cattle spread across Karamoja region. Serum was extracted and screened for antibodies against Mycoplasma mycoides subsp. mycoides [Mmm] using the competitive enzyme linked immunosorbent assay [cELISA]. Results A quarter [25.4%; 95% CI: 23.7–27.3] of the screened cattle [n = 2,300] were sero-positive for Mmm. Amudat and Kaabong districts recorded the lowest [12.3%] and highest [30.7%] Mmm seroprevalence respectively. Increasing age, overnight stay in cattle kraals and location [certain districts, villages, herds and sub counties] of the cattle herds, the factors that promote animal commingling, were the most significant risk factors of seroconversion with Mmm. Conclusion Results from this study indicated a higher seroprevalence of Mmm in Karamoja region cattle herds. This could be due to the increased frequency of CBPP outbreaks in recent years. To be effective, CBPP vaccination programs should target high risk herds along the international borders and other hotspot areas [e.g., parishes or sub counties] where cattle commingling is high.

Background: Tick-borne encephalitis (TBE) is a severe human neuroinfection caused by TBE virus (TBEV). TBEV is transmitted by tick bites and by the consumption of unpasteurized dairy products from infected asymptomatic ruminants. In France, several food-borne transmission events have been reported since 2020, raising the question of the level of exposure of domestic ungulates to TBEV. In this study, our objectives were (i) to estimate TBEV seroprevalence and quantify antibodies titres in cattle in the historical endemic area of TBEV in France using the micro virus neutralisation test (MNT) and (ii) to compare the performance of two veterinary cELISA kits with MNT for detecting anti-TBEV antibodies in cattle in various epidemiological contexts. A total of 344 cattle sera from four grid cells of 100 km² in Alsace-Lorraine (endemic region) and 84 from western France, assumed to be TBEV-free, were investigated. Results: In Alsace-Lorraine, cattle were exposed to the virus with an overall estimated seroprevalence of 57.6% (95% CI: 52.1–62.8%, n = 344), varying locally from 29.9% (95% CI: 21.0–40.0%) to 92.1% (95% CI: 84.5–96.8%). Seroprevalence did not increase with age, with one- to three-year-old cattle being as highly exposed as older ones, suggesting a short-life duration of antibodies. The proportion of sera with MNT titres lower than 1:40 per grid cell decreased with increased seroprevalence. Both cELISA kits showed high specificity (> 90%) and low sensitivity (less than 78.1%) compared with MNT. Sensitivity was lower for sera with neutralising antibodies titres below 1:40, suggesting that sensitivity of these tests varied with local virus circulation intensity. Conclusions: Our results highlight that cattle were highly exposed to TBEV. Screening strategy and serological tests should be carefully chosen according to the purpose of the serological study and with regard to the limitations of each method. [ABSTRACT FROM AUTHOR]

Background: Contagious bovine pleuropneumonia [CBPP] is a transboundary animal disease of cattle caused by Mycoplasma mycoides subsp. mycoides [Mmm]. CBPP causes severe economic losses to livestock producers in sub-Saharan Africa mainly due to high mortality, morbidity, reduction in productivity as well as livestock trade restrictions. This study aimed at determining seroprevalence of Mmm in cattle from Karamoja region, north-eastern Uganda; data that are required to design and implement risk based CBPP control program. Methods: We randomly collected blood samples from 2,300 cattle spread across Karamoja region. Serum was extracted and screened for antibodies against Mycoplasma mycoides subsp. mycoides [Mmm] using the competitive enzyme linked immunosorbent assay [cELISA]. Results: A quarter [25.4%; 95% CI: 23.7–27.3] of the screened cattle [n = 2,300] were sero-positive for Mmm. Amudat and Kaabong districts recorded the lowest [12.3%] and highest [30.7%] Mmm seroprevalence respectively. Increasing age, overnight stay in cattle kraals and location [certain districts, villages, herds and sub counties] of the cattle herds, the factors that promote animal commingling, were the most significant risk factors of seroconversion with Mmm. Conclusion: Results from this study indicated a higher seroprevalence of Mmm in Karamoja region cattle herds. This could be due to the increased frequency of CBPP outbreaks in recent years. To be effective, CBPP vaccination programs should target high risk herds along the international borders and other hotspot areas [e.g., parishes or sub counties] where cattle commingling is high. [ABSTRACT FROM AUTHOR]

Peste des petits ruminants (PPR) is a highly contagious and devastating viral disease of goats and sheep caused by Peste des petits ruminants virus (PPRV). Despite of its impact on the livelihood of rural African communities, insufficient epidemiological information hampers the implementation of effective PPR control strategies. This study was carried out to determine seroprevalence and risk factors for PPR infection in sheep and goats in Longido, Simanjiro, Ngorongoro, Monduli, Kongwa and Mlele districts using a competitive Enzyme--Linked Immunosorbent Assay and questionnaire to detect PPR antibodies, and to collect information related to the potential risk factors for PPR respectively. A total of 583 serum samples from sheep (n=248), and goats (n=335), and 40 households were involved in questionnare. This study confirmed presence of antibodies to PPRV in sheep and goats in all districts studied; and identified management systems related to movement control to be an important risk factor for the spread of PPR. The results suggest that controlling animal movement and limiting interaction with wildlife could PPR transmission. [ABSTRACT FROM AUTHOR]

Introduction: To determine clinico-pathology caused by Mycoplasma mycoides subspecies capri (PG3) in goats of different age groups and antibody titers. Methods: Twenty-one goats of three age groups viz; 1, 2 and 3 years with 7 goats of each age. Goats were divided in four groups i-e; G1, G2, G3 each contain 5 goats of each age while G4 comprise 6 goats; 2 from each group. Goats in G1, G2 and G3 were infected with Mycoplasma mycoides (PG3) dose (1x107 CFU/ml/kg) whereas, G4 was kept as control. Clinical examinations were recorded at 12-h intervals. The blood samples collected were tested through cELISA and the specimen of trachea, lungs, kidney, and liver were collected at the end of experiment for gross and histopathology. Results: Temperature, respiratory rate, pulse rate nasal discharge, coughing and lacrimation were noted higher in G1 compared to G2 and G3 groups. Gross pathology showed severe multifocal and diffused necrosis G1 compared to G2 and G3 groups. Histopathology showed sloughing of tracheal mucosa in all groups while hypertrophic secretary glands in G1. Lungs showed emphysema in all groups except G4. Kidneys showed glomerulonephritis while Liver showed congestion and hyperemia in all groups. cELISA, revealed the antibody titers rose from 1st to 3rd week post infection afterwards, reduced slowly. Antibody titers were higher in G1 compared to G2 and G3 groups. Conclusion: Mycoplasma mycoides subspecies capri (PG3) can cause infection in goats of all ages, yet the infection is more severe in young animals compared to old. [ABSTRACT FROM AUTHOR]

The peste des petits ruminant (PPR) virus is a transboundary virus found in small domestic ruminants that causes high morbidity and mortality in naive herds. PPR can be effectively controlled and eradicated by vaccinating small domestic ruminants with a live-attenuated peste des petits ruminant virus (PPRV) vaccine, which provides long-lasting immunity. We studied the potency and safety of a live-attenuated vaccine in goats by detecting their cellular and humoral immune responses. Six goats were subcutaneously vaccinated with a live-attenuated PPRV vaccine according to the manufacturer's instructions, and two goats were kept in contact. Following vaccination, the goats were monitored daily, and we recorded their body temperature and clinical score. Heparinized blood and serum were collected for a serological analysis, and swab samples and EDTA blood were collected to detect the PPRV genome. The safety of the used PPRV vaccine was confirmed by the absence of PPR-related clinical signs, a negative pen-side test, a low virus genome load as detected with RT-qPCR on the vaccinated goats, and the lack horizontal transmission between the in-contact goats. The strong humoral and cellular immune responses detected in the vaccinated goats showed that the live-attenuated PPRV vaccine has a strong potency in goats. Therefore, live-attenuated vaccines against PPR can be used to control and eradicate PRR. [ABSTRACT FROM AUTHOR]

Schmallenberg virus (SBV) is an emerging pathogen of sheep. It is transmitted through arthropods vertically from dams to fetuses through the placenta. As a result, it can cause prenatal defects that can result in abortion or death of the fetus, as well as sometimes the death of mothers due to complications following an abortion. The study was conducted on 92 sera of local breed sheep (67 aborted ewes and 25 rams). All sera were tested for Schmallenbergvirusantibodies using a competitive enzyme-linked immunosorbent kit. Results indicated that the overall prevalence of the Schmallenbergviruswas 79.3 %. The highest percentages of SBV seropositive animals were found in Mosul city, Wana, and Tel Usqouf at 100%, whereas Badoush reported the lowest percentage at 40%. Additionally, there was a significant difference between the percentage of aborted ewes 85.07% and rams 64%. The most frequent infection type was the severe one 55,43%, followed by the moderate 18.47% and mild 5.43% types. In conclusion, the findings highlight the existence of a novel virus that significantly impacts ovine abortions. So, these preliminary data can guide livestock producers in designing appropriate prevention and control measures.

Abstract Background Several reports suggest a further spread of besnoitiosis to countries in which Besnoitia besnoiti-infected bovine herds have not been noticed yet. Cattle infected without clinical signs may represent reservoirs. Serological analyses in affected herds or animals from endemic regions are necessary to identify subclinical or inapparent infections and stop transmission to naïve animals or herds. The Monoscreen AbELISA Besnoitia besnoiti (BIO K 466) is based on a previously published in-house competitive ELISA, the Bb-cELISA1, but has a different test architecture. The present study aimed to use sera from a previous evaluation of Bb-cELISA1 to assess whether BIO K466 shows identical results. In addition, further well-characterized positive and negative samples were analysed to estimate diagnostic sensitivity and specificity. Methods A first set of sera consisted of a total of 305 bovine sera, collected from German herds infected by B. besnoiti, Neospora caninum or Sarcocystis spp. Sera had been characterized by reference serological tests (i.e. immunoblot, immunofluorescence antibody test and an in-house indirect ELISA). A second set consisted of 200 confirmed B. besnoiti-positive sera from French herds. Negative cattle sera (n = 624) originated from Norway and The Netherlands, countries in which bovine besnoitiosis has not been reported yet. Results Using the first set of sera, the BIO K466 showed an estimated diagnostic sensitivity of 97.9% (95% CI: 91.9%–99.6) and a diagnostic specificity of 99.5% (95% CI: 96.9%–100%) relative to reference serological tests. A direct comparison of the results revealed an almost perfect agreement between the results of the in-house Bb-cELISA1 and the commercialized version (kappa 0.98; 95% CI: 0.95–1). The validation using positive bovine sera from France and negative sera from other European countries revealed a diagnostic sensitivity of 97.5% (95% CI: 93.9%–99.1%) and specificity of 99.5% (95% CI: 98.5%–99.9%). Conclusion In conclusion, BIO K 466 appears to be a suitable tool to diagnose bovine besnoitiosis, but needs further validation especially in cases of inconclusive, suspected false-positive or -negative results in other serological tests. Graphic abstract

ABSTRACT Hepatitis E virus (HEV) is a zoonotic pathogen that is widespread worldwide. At present, most enzyme-linked immunosorbent assay (ELISA) kits only detect antibodies against human HEV. In this study, a nanobody-horseradish peroxidase (HRP) fusion protein-based competitive ELISA (cELISA) with more convenience and spectral characteristics for HEV antibody detection was developed and used to detect HEV IgG in various species. First, 6 anti-swine HEV capsid protein nanobodies were screened using phage display technology from an immunized Bactrian camel. Then, HEV-Nb67-HRP fusions were expressed and used as a probe for developing a cELISA. The cutoff value of the cELISA was 17.8%, and there was no cross-reaction with other anti-swine virus antibodies, suggesting that the cELISA had good specificity. The intra-assay and interassay coefficients of variation (CVs) were 1.33 to 5.06% and 1.52 to 6.84%, respectively. The cELISA and Western blot showed a higher coincidence rate (97.14%, kappa value = 0.927) than cELISA and indirect ELISA (95.00%, kappa value = 0.876) in clinical swine serum samples. Finally, the seroprevalence of HEV IgG in humans, pigs, rabbits, cows, and goats was 30.67%, 19.26%, 8.75%, 27.59%, and 18.08%, respectively, suggesting that cELISA may have a broader scale for mammalian HEV antibody detection. These results suggest that the newly developed cELISA was rapid, low-cost, reliable, and useful for the serological evaluation of current HEV. IMPORTANCE HEV is thought to be a zoonotic infection and is widespread worldwide; it is beneficial to establish a more convenient and spectral method for HEV antibody detection. In this study, a convenient, time-saving, reproducible, highly sensitive, specific, and novel nanobody-based cELISA was developed and can be used to detect IgG antibodies against mammalian HEV. It provides a new technique for serological evaluation and ELISA-based diagnosis of HEV infection.

Abstract Background African swine fever virus (ASFV) is a highly contagious hemorrhagic disease and often lethal, which has significant economic consequences for the swine industry. Due to lacking of commercial vaccine, the prevention and control of ASF largely depend on early large-scale detection and screening. So far, the commercial ELISA kits have a long operation time and are expensive, making it difficult to achieve large-scale clinical applications. Nanobodies are single-domain antibodies produced by camelid animals, and have unique advantages such as smaller molecular weight, easy genetic engineering modification and low-costing of mass production, thus exhibiting good application prospects. Results The present study developed a new method for detection of ASFV specific antibodies using nanobody-horseradish peroxidase (Nb-HRP) fusion proteins as probe. By using camel immunization, phage library construction and phage display technology, five nanobodies against K205R protein were screened. Then, Nb-HRP fusion proteins were produced using genetic modification technology. Based on the Nb-HRP fusion protein as specific antibodies against K205R protein, a new type of cELISA was established to detect ASFV antibodies in pig serum. The cut-off value of the cELISA was 34.8%, and its sensitivity, specificity, and reproducibility were good. Furthermore, the developed cELISA exhibited 99.3% agreement rate with the commercial available ELISA kit (kappa value = 0.98). Conclusions The developed cELISA method has the advantages of simple operation, rapid and low-costing, and can be used for monitoring of ASFV infection in pigs, thus providing a new method for the prevention and control of ASF.

Simple Summary: Brucellosis is a neglected, bacterial zoonotic disease that affects domesticated animals and people. Infection in cattle is caused by Brucella abortus which causes nonspecific clinical signs in female cattle including lowered milk production, infertility, and abortion in the last trimester. To determine the prevalence and the risk factors associated with brucellosis exposure, we conducted a cross-sectional study of smallholder dairy cattle in six regions of Tanzania, between July 2019 and October 2020. A total of 2048 dairy cattle blood samples were collected and tested for the presence of anti-Brucella antibodies. An overall seroprevalence of 2.39% was found with the highest seroprevalence in the Njombe Region (15.5%). The risk factors that were identified to be significantly associated with brucellosis seropositivity were having goats around dairy cattle and a history of abortion within a farm. The study findings suggest that brucellosis is still present in smallholder dairy cattle at varying levels among the studied regions. Education of smallholder dairy keepers is required regarding the disease, as well as risk and control measures for the disease. A One Health approach is required to study the role of small ruminants in the spread of the disease and to evaluate the public health risk to smallholder dairy farmers, especially in the Njombe Region. Bovine brucellosis is a bacterial zoonoses caused by Brucella abortus. We conducted a cross-sectional study to determine brucellosis seroprevalence and risk factors among smallholder dairy cattle across six regions in Tanzania. We sampled 2048 dairy cattle on 1374 farms between July 2019 and October 2020. Sera were tested for the presence of anti-Brucella antibodies using a competitive enzyme-linked immunosorbent assay. Seroprevalence was calculated at different administrative scales, and spatial tests were used to detect disease hotspots. A generalized mixed-effects regression model was built to explore the relationships among Brucella serostatus, animals, and farm management factors. Seroprevalence was 2.39% (49/2048 cattle, 95% CI 1.7–3.1) across the study area and the Njombe Region represented the highest percentage with 15.5% (95% CI 11.0–22.0). Moreover, hotspots were detected in the Njombe and Kilimanjaro Regions. Mixed-effects models showed that having goats (OR 3.02, 95% C 1.22–7.46) and abortion history (OR 4.91, 95% CI 1.43–16.9) were significant risk factors for brucellosis. Education of dairy farmers regarding the clinical signs, transmission routes, and control measures for brucellosis is advised. A One Health approach is required to study the role of small ruminants in cattle brucellosis and the status of brucellosis in dairy farmers in the Njombe and Kilimanjaro Regions. [ABSTRACT FROM AUTHOR]

Anaplasmosis, a tick-borne disease affecting cattle worldwide caused by Anaplasma spp. (Rickettsiales: Anaplasmataceae), is associated with considerable economic losses to dairy and beef industries. Data about exposure to Anaplasma spp. infection of cattle in Romania is limited. Therefore, a cross-sectional serological study was conducted to preliminary investigate the serological status on Anaplasma spp. infection in Romanian catlle. For this, 182 catlle originated from three counties in Northeastern and Southeastern Romania were included in the study. Cattle serum samples were tested with a competitive enzyme linked immunosorbent assay (cELISA) detecting an epitope of major surface protein 5 (MSP5) of Anaplasma spp. The results were expressed as percentage (%) of inhibition (I) calculated according to the indications of the manufactures; samples with I ≥30% were considered positive. Subsequently, the overall seropositivity was 42.3% (95% CI: 35.03 - 49.84); significant difference of seroprevalence values (p< 0.05) were registered according to the cattle originating county, ranging from 22.2% (95% CI: 13.26-33.57) to 72.0% (95% CI: 57.50-83.77). These findings emphasize on the need for further studies to identify the species and associated risk factors for Anaplasma inffection of cattle in Romania, as base for developing control strategies. [ABSTRACT FROM AUTHOR]

Background: Several reports suggest a further spread of besnoitiosis to countries in which Besnoitia besnoiti-infected bovine herds have not been noticed yet. Cattle infected without clinical signs may represent reservoirs. Serological analyses in affected herds or animals from endemic regions are necessary to identify subclinical or inapparent infections and stop transmission to naïve animals or herds. The Monoscreen AbELISA Besnoitia besnoiti (BIO K 466) is based on a previously published in-house competitive ELISA, the Bb-cELISA1, but has a different test architecture. The present study aimed to use sera from a previous evaluation of Bb-cELISA1 to assess whether BIO K466 shows identical results. In addition, further well-characterized positive and negative samples were analysed to estimate diagnostic sensitivity and specificity. Methods: A first set of sera consisted of a total of 305 bovine sera, collected from German herds infected by B. besnoiti, Neospora caninum or Sarcocystis spp. Sera had been characterized by reference serological tests (i.e. immunoblot, immunofluorescence antibody test and an in-house indirect ELISA). A second set consisted of 200 confirmed B. besnoiti-positive sera from French herds. Negative cattle sera (n = 624) originated from Norway and The Netherlands, countries in which bovine besnoitiosis has not been reported yet. Results: Using the first set of sera, the BIO K466 showed an estimated diagnostic sensitivity of 97.9% (95% CI: 91.9%–99.6) and a diagnostic specificity of 99.5% (95% CI: 96.9%–100%) relative to reference serological tests. A direct comparison of the results revealed an almost perfect agreement between the results of the in-house Bb-cELISA1 and the commercialized version (kappa 0.98; 95% CI: 0.95–1). The validation using positive bovine sera from France and negative sera from other European countries revealed a diagnostic sensitivity of 97.5% (95% CI: 93.9%–99.1%) and specificity of 99.5% (95% CI: 98.5%–99.9%). Conclusion: In conclusion, BIO K 466 appears to be a suitable tool to diagnose bovine besnoitiosis, but needs further validation especially in cases of inconclusive, suspected false-positive or -negative results in other serological tests. [ABSTRACT FROM AUTHOR]

H7 avian influenza viruses (AIVs) normally circulated among birds before. From 1996 to 2012, human infections with H7 AIVs (H7N2, H7N3, and H7N7) were reported in Canada, Italy, Mexico, the Netherlands, the United Kingdom and the USA. Until March 2013, human infections with H7N9 AIVs were reported in China. Since then, H7N9 AIVs have continued to circulate in both humans and birds. Therefore, the detection of antibodies against the H7 subtype of AIVs has become an important topic. In this study, a competitive enzyme-linked immunosorbent assay (cELISA) method for the detection of antibody against H7 AIVs was established. The optimal concentration of antigen coating was 5 μg mL−1, serum dilution was 1/10, and enzyme-labeled antibody was 1/3 000. To determine the cut-off value of cELISA, percent inhibition (PI) was determined by using receiver operating characteristic (ROC) curve analysis in 178 AIVs negative samples and 368 AIVs positive serum samples (n=546). When PI was set at 40%, the specificity and sensitivity of cELISA were 99.4 and 98.9%, respectively. This method could detect the antibodies against H7Nx (N1–N4, N7–N9) AIVs, and showed no reaction with AIVs of H1–H6 and H8–H15 subtypes or common avian viruses such as Newcastle disease virus (NDV), Infectious bronchitis virus (IBV) and Infectious bursal disease virus (IBDV), exhibiting good specificity. This method showed a coincidence rate of 98.56% with hemagglutinin inhibition (HI) test. And the repeatability experiment revealed that the coefficients of variation (CV) of intra- and inter-batch repetition were all less than 12%. The data indicated that the cELISA antibody-detection method established in this study provided a simple and accurate technical support for the detection of a large number of antibody samples of H7-AIV.

Background: African swine fever virus (ASFV) is a highly contagious hemorrhagic disease and often lethal, which has significant economic consequences for the swine industry. Due to lacking of commercial vaccine, the prevention and control of ASF largely depend on early large-scale detection and screening. So far, the commercial ELISA kits have a long operation time and are expensive, making it difficult to achieve large-scale clinical applications. Nanobodies are single-domain antibodies produced by camelid animals, and have unique advantages such as smaller molecular weight, easy genetic engineering modification and low-costing of mass production, thus exhibiting good application prospects. Results: The present study developed a new method for detection of ASFV specific antibodies using nanobody-horseradish peroxidase (Nb-HRP) fusion proteins as probe. By using camel immunization, phage library construction and phage display technology, five nanobodies against K205R protein were screened. Then, Nb-HRP fusion proteins were produced using genetic modification technology. Based on the Nb-HRP fusion protein as specific antibodies against K205R protein, a new type of cELISA was established to detect ASFV antibodies in pig serum. The cut-off value of the cELISA was 34.8%, and its sensitivity, specificity, and reproducibility were good. Furthermore, the developed cELISA exhibited 99.3% agreement rate with the commercial available ELISA kit (kappa value = 0.98). Conclusions: The developed cELISA method has the advantages of simple operation, rapid and low-costing, and can be used for monitoring of ASFV infection in pigs, thus providing a new method for the prevention and control of ASF. [ABSTRACT FROM AUTHOR]

African swine fever (ASF), which is caused by the ASF virus (ASFV), is a highly contagious hemorrhagic disease that causes high mortality to domestic porcine and wild boars and brings huge economic losses to world swine industry. Due to the lack of an effective vaccine, the control of ASF must depend on early, efficient, and cost-effective detection and strict control and elimination strategies. Traditional serological testing methods are generally associated with high testing costs, complex operations, and high technical requirements. As a promising alternative diagnostic tool to traditional antibodies, nanobodies (Nb) have the advantages of simpler and faster generation, good stability and solubility, and high affinity and specificity, although the system is dependent on the immunization of Bactrian camels to obtain the specific VHH library of the target protein. The application of Nbs in the detection of ASFV antibodies has not yet been reported yet. Using a phage display technology, one Nb against the ASFV p54 protein that exhibited high specificity and affinity, Nb8, was successfully screened. A HEK293T cell line stably expressing Nb8-horseradish peroxidase (HRP) fusion protein was established using the lentiviral expression system. Following the optimization of the reaction conditions, the Nb8-HRP fusion protein was successfully used to establish a competitive enzyme-linked immunosorbent assay (cELISA) to detect ASFV-specific antibodies in pig serum, for the first time. There was no cross-reaction with healthy pig serum, porcine pseudorabies virus (PRV), porcine reproductive and respiratory syndrome virus (PRRSV), classical swine fever virus (CSFV), porcine epidemic diarrhea virus (PEDV), and classical swine fever virus (CSFV) positive sera. The optimal cut-off value for the cELISA by ROC analysis was 52.5%. A total of 209 serum samples were tested using the developed cELISA and a commercial ELISA kit. The results showed that the relative specificity of the cELISA was 98.97%, and the relative sensitivity of the cELISA was 93.3%, with the percent agreement between the two ELISA methods being 98.56%. In conclusion, a specific, sensitive, and repeatable cELISA was successfully developed based on the Nb8 as a probe, providing a promising method for the detection of anti-ASFV antibodies in clinical pig serum. Key points: • We successfully screened a specific, high affinity nanobody against ASFV p54 protein. • We establish a method for continuous and stable expression of Nb-HRP fusion protein using a lentiviral packaging system. • We establish a nanobody cELISA detection method that can monitor an ASF infection. [ABSTRACT FROM AUTHOR]

Equine piroplasmosis (EP) is an endemic tick-borne disease found in most countries around the world. It affects all species of Equidae, and it is caused by Theileria equi, Babesia caballi and T. haneyi. The research herein is the second study on the prevalence of piroplasms in Romania conducted in the past two decades. The aim of this study was to assess the seroprevalence of anti-Theileria equi antibodies and the geographical distribution of this disease in the southwest, west, and northwest regions of Romania in order to obtain a more thorough understanding of the parasitological status of horses in this country. This study included 522 apparently healthy, mixed-breed horses from three different counties. The serum samples were analysed using the cELISA Theileria equi Antibody Test Kit. The overall seroprevalence rate was 12.84%. From the total number of positive horses, 13.96% were females and 11.21% were males. Based on the distribution of positive cases into age groups, the following values were obtained: 0–60 months: 16.26%, 60–180 months: 10.03%, and >180 months: 15.83%. There was no statistically significant difference between samples, based on age or gender. The positivity percentage in the localities included in the study ranged from 8.33 to 100%. In the population under study, the seroprevalence rate was high, indicating a possible exposure risk in this area of Romania, which could have severe effects on equids in the case of clinical manifestations of the disease. EP represents a serious threat for equine health in Romania; therefore, close and continuous monitoring of the situation is required. [ABSTRACT FROM AUTHOR]

Anaplasma spp. is a group of intra-erythrocytic bacteria that various species of ticks can transmit. Information regarding the prevalence of infections in sheep and goats caused by this group of organisms is scarce in Saudi Arabia. The present study was carried out during 2020–2021 to examine the prevalence of anaplasmosis among sheep and goats in two cities in western Saudi Arabia. The study included samples from 177 sheep (77 from Madina, 100 from Tabuk) and 226 goats (123 from Madina, 103 from Tabuk). The samples were investigated using direct microscopy method as well as a competitive Enzyme-Linked Immuno-Assay (cELISA) for the detection of anti-Anaplasma spp. antibodies. A total of 93 (23.1%) of the samples were positive on direct microscopy, whereas 84 (20.7%) were positive on cELISA. Of those samples positive on direct microscopy, 44 (19.5%) were from goats while 49 (27.7%) were from sheep. Of the positive samples on cELISA; 38 (17.0%) were from goats, and 46 (26.0%) were from sheep. A significant difference in the prevalence of anaplasmosis was reported (p 0.05). However, there was a significant difference in the prevalence of anaplasmosis in older goats (>2 years) and in the summer compared to the winter in samples collected from Tabuk (p

Schmallenberg virus is an emergent disease which infect cattle, sheep and goats which cause loss of condition, diarrhea, and abortion in pregnant animals, so this study was conducted to detect the antibodies in imported calves by using cELISA, so 400 blood samples was collected calves in different ages and healthy status in a period between October 2018 to September 2019 in Nineveh province, the investigation of specific antibodies was done by competitive Enzyme Linked Immuno Sorbent Assay, the results showed that the prevalence of Schmallenberg virus in imported calves was 21% (84 positive from 400 samples), high prevalence of infection in the animals more than 6 months to 1 year old 11.5% when compare with animals less than 6 months of age 9.5%, high incidence of infection in animals suffer from various clinical signs 17% and the healthy apparent animals recorded low prevalence of infection 4%, samples which collected in spring months recorded high prevalence of infection 7.5% while the lowest prevalence of infection with the virus recorded in winter months 2%, with significant variance in spring and summer months compared with other seasons, in conclusion this study was conducted that schmallenberg virus is newly emerge in Iraq and this study is firstly recorded this virus in cattle in Iraq.

Abstract Background Bluetongue (BT) is a vector-borne viral disease of ruminant and camelid species which is transmitted by Culicoides spp. The causative agent of BT is bluetongue virus (BTV) that belongs to genus Orbivirus of the family Reoviridae. The clinical disease is seen mainly in sheep but mostly sub-clinical infections of BT are seen in cattle, goats and camelids. The clinical reaction of camels to infection is usually not apparent. The disease is notifiable to the World Organization for Animal Health (OIE), causing great economic losses due to decreased trade and high mortality and morbidity rates associated with bluetongue outbreaks. The objective of this study was to investigate the seroprevalence of BTV in camels in Kassala State, Eastern Sudan and to identify the potential risk factors associated with the infection. A cross sectional study using a structured questionnaire survey was conducted during 2015–2016. A total of 210 serum samples were collected randomly from camels from 8 localities of Kassala State. The serum samples were screened for the presence of BTV specific immunoglobulin (IgG) antibodies using a competitive enzyme-linked immunosorbent assay (cELISA). Results Seropositivity to BTV IgG was detected in 165 of 210 camels’ sera accounting for a prevalence of 78.6%. Potential risk factors to BTV infection were associated with sex (OR = 0.061, p-value = 0.001) and seasonal river as water source for drinking (OR = 32.257, p-value = 0.0108). Conclusions Sex and seasonal river as water source for drinking were considered as potential risk factors for seropositivity to BTV in camels. The high prevalence of BTV in camels in Kassala State, Eastern Sudan, necessitates further epidemiological studies of BTV infection in camels and other ruminant species to better be able to control BT disease in this region.

Equine piroplasmosis (EP) is a tick-borne disease of horses, donkeys and wild equids. Chronically infected animals, which often only show reduced performance or are clinically unremarkable, constitute a potential risk for parasite spread. In Europe, currently no transboundary preventive measures for identifying infected animals or preventing their cross-border transportation are in place. In Austria, as in the neighbouring countries of Switzerland and Germany, EP has only been detected sporadically. By contrast, Austria’s eastern neighbour, Hungary, is considered endemic for EP. The geographic vicinity to Hungary and a recently described autochthonous case of EP induced the present pilot study on the serological status of horses in eastern Austria. Serum samples of 173 clinically healthy horses of different age and sex from 43 different locations in Vienna and Lower Austria, without known stays abroad within the last 12 months, were examined by cELISA. Sampling had been conducted during a prevalence study on West Nile infections in 2017. Four horses (2.3 %) aged 7–28 years from three different locations were positive for anti-T. equi antibodies. Two of the positive horses had a Hungarian passport, and two horses an Austrian passport. One of the horses with an Austrian passport, however, was born in Romania. No horse was positive for anti-B. caballi-antibodies. The results of this study show that animals infected with T. equi occur in the east of Austria. Equine practitioners should consider EP as a possible cause of febrile diseases and haemolytic anaemia, but also be aware of chronic or subclinical courses of infection and the risk of parasite transmission from inapparent carriers. In addition, EP should be monitored in areas with known occurrence of Dermacentor as a vector-competent tick genus. [ABSTRACT FROM AUTHOR]

Anaplasmosis is an infectious disease affecting goats and other ruminants. Our goal was to estimate the prevalence of Anaplasma spp. infections, and to identify different tick species in goa ts along with their associated risk factors. The epidemiology of Anaplasma spp. (A. ovis/A. marginale) was investigated from April to September 2016 in dairy goats from three different agro-ecological areas in the northeast region of Algeria (Guelma and El-Taref). We tested 182 goat sera using a MSP5 competitive ELISA (cELISA) test for IgG antibody screening, and by microscopic examination using the Giemsa (May-Greunwald-Giemsa) stain of 128 blood smears to detect intraerythrocytic inclusions bodies. The results demonstrated a total infection rate of 78.02 % and 42.18 % using cELISA and MGG, respectively. Only two species of ticks collected were identified, i. e. Rhipicephalus sanguineus (85.39 %) and Rhipicephalus bursa (14.60 %). Our study revealed that factors, such as the season, the type of farming, the hygienic status and the climatic conditions of the studied areas were found to be significantly associated with the tick infestation of goats (P < 0.05). The Anaplasma spp. infection prevalence was found to be highly dependent on the climatic conditions, the season (P < 0.05) and most closely related to the type of breeding and herd management (P = 0.000).

Bovine brucellosis is a bacterial zoonoses caused by Brucella abortus. We conducted a cross-sectional study to determine brucellosis seroprevalence and risk factors among smallholder dairy cattle across six regions in Tanzania. We sampled 2048 dairy cattle on 1374 farms between July 2019 and October 2020. Sera were tested for the presence of anti-Brucella antibodies using a competitive enzyme-linked immunosorbent assay. Seroprevalence was calculated at different administrative scales, and spatial tests were used to detect disease hotspots. A generalized mixed-effects regression model was built to explore the relationships among Brucella serostatus, animals, and farm management factors. Seroprevalence was 2.39% (49/2048 cattle, 95% CI 1.7–3.1) across the study area and the Njombe Region represented the highest percentage with 15.5% (95% CI 11.0–22.0). Moreover, hotspots were detected in the Njombe and Kilimanjaro Regions. Mixed-effects models showed that having goats (OR 3.02, 95% C 1.22–7.46) and abortion history (OR 4.91, 95% CI 1.43–16.9) were significant risk factors for brucellosis. Education of dairy farmers regarding the clinical signs, transmission routes, and control measures for brucellosis is advised. A One Health approach is required to study the role of small ruminants in cattle brucellosis and the status of brucellosis in dairy farmers in the Njombe and Kilimanjaro Regions.

Background: Theileria equi and Babesia caballi cause equine piroplasmosis (EP), one of the most important tick‐borne diseases of horses due to its high negative impact to the equine industry. Although infections with these parasites have been reported for decades in Spain, epidemiological studies have only been carried out in certain regions. Objectives: To determine the (sero)prevalence of these parasites in asymptomatic horses nationwide in Spain and to identify potential individual and environmental factors associated with seropositivity to EP. Study design: Sample size was calculated according to the horses registered in Spain in 2013 and by autonomous community using a random stratified sampling. A questionnaire was used to collect data on factors associated with EP seropositivity. Methods: Serological (cELISA and complement fixation test) and molecular (real‐time PCR) analyses were carried out in 740 horses. Risk factors were identified computing two independent logistic regression models with the collated data. Results: Antibodies against EP were detected in 42.9% (95% CI 39.4‐46.5) of horses, whereas 30.3% (95% CI 27.0‐33.6) were EP positive by PCR. A substantial strength of agreement (k = 0.721) was estimated between serological tests. Exposure to T. equi was significantly higher than to B. caballi and the highest (sero)prevalence was detected in the northern communities. Increasing horse age, presence of ticks and contact with cows were factors related to EP seropositivity in the horses, whereas tetanus vaccination and fairs attendance were associated with lower seropositivity. Conclusions: Almost half of the horses residing in Spain had antibodies against EP or circulating parasitaemia. Appropriate prevention measures and implementation of a EP surveillance programme should be considered in order to reduce and control the infection. [ABSTRACT FROM AUTHOR]

Objectives: The study was undertaken with the objectives to perform seromonitoring of Peste des Petits Ruminants (PPR) antibodies in goats vaccinated with PPR vaccine and molecular character¬ization of PPR virus (PPRV) from field cases in Bangladesh. Materials and Methods: Seromonitoring work was conducted in Char Kalibari, Mymensingh Sadar, Mymensingh. For this, a total of 50 goats were randomly selected and were divided into two groups; vaccinated (Group A; n = 25) and non-vaccinated (Group B; n = 25). The goats of both groups were again sub-divided into four age groups; (i) 06 months (n = 5), (ii) 612 months (n = 5), (iii) 1224 months (n = 10), and (iv) >24 months (n = 5). Blood samples were collected on Day-0 and after 21 days of post-vaccination (DPV), and the sera were prepared. The sera were examined for the presence of antibodies against PPRV by competitive enzyme-linked immunosorbent assay. For molecular characterization, nasal swabs (n = 10) were collected from PPR infected goats in Jessore during PPR outbreak (February 2016). The causative agent, PPRV isolated from field cases were confirmed by N gene based on reverse transcription polymerase chain reaction (RT-PCR), followed by sequencing, phylogenetic analysis, and multiple sequence alignment analyses. Results: In the case of seromonitoring, the results revealed that before vaccination (at Day-0), overall, 44% (n = 22/50) goats were seropositive for PPRV. In Group A, 48% (n = 12/25) goats were seropositive, but after 21 DPV, 96% (n = 24/25) goats become seropositive. On the other hand, in Group B, 40% (n = 10/25) and 16% (n = 04/25) seropositive goats found at Day-0 and after 21 DPV, respectively, indicating that the antibody titer was increasing after vaccination and decreasing in convalescent goats. Out of 10 nasal swab samples, 40% (n = 4/10) was confirmed by RT-PCR targeting nucleocapsid (N gene). Phylogenetically, our isolate (KY039156/PPRV/BDG/Jes/2016) was similar to the other strains of PPRV under lineage IV. However, there was a unique amino acid substitution, where glycine (G) was recorded in place of arginine (R). The strain is closely related with other Chinese or Indian strains. The nucleotide sequence homology by NCBI BLAST search of the isolated strain ranged from 95% to 99% with other strains circulating in Bangladesh. Conclusion: The PPRV is prevailing in the Mymensingh and Jessore regions of Bangladesh. Effective control of PPR in goats may depend on vaccination with PPR vaccine. Molecular characterization of PPRV in Jessore reveals that the virus is differing from the strain prevalent in other regions of Bangladesh and the world. [J Adv Vet Anim Res 2019; 6(3.000): 416-424]

Equine piroplasmosis (EP) is an endemic tick-borne disease found in most countries around the world. It affects all species of Equidae, and it is caused by Theileria equi, Babesia caballi and T. haneyi. The research herein is the second study on the prevalence of piroplasms in Romania conducted in the past two decades. The aim of this study was to assess the seroprevalence of anti-Theileria equi antibodies and the geographical distribution of this disease in the southwest, west, and northwest regions of Romania in order to obtain a more thorough understanding of the parasitological status of horses in this country. This study included 522 apparently healthy, mixed-breed horses from three different counties. The serum samples were analysed using the cELISA Theileria equi Antibody Test Kit. The overall seroprevalence rate was 12.84%. From the total number of positive horses, 13.96% were females and 11.21% were males. Based on the distribution of positive cases into age groups, the following values were obtained: 0–60 months: 16.26%, 60–180 months: 10.03%, and >180 months: 15.83%. There was no statistically significant difference between samples, based on age or gender. The positivity percentage in the localities included in the study ranged from 8.33 to 100%. In the population under study, the seroprevalence rate was high, indicating a possible exposure risk in this area of Romania, which could have severe effects on equids in the case of clinical manifestations of the disease. EP represents a serious threat for equine health in Romania; therefore, close and continuous monitoring of the situation is required.

Contagious caprine pleuropneumonia (CCPP) is an economically important and potentially fatal disease of small ruminants caused by Mycoplasma capricolum subsp. capripneumoniae (Mccp). We designed this cross-sectional study to investigate the seroepidemiology of CCPP in the small ruminants of Oman. For this purpose, we sampled a total of 4015 small ruminants (2119 goats and 1896 sheep) from 510 flocks belonging to different governorates of Oman. A commercial competitive enzyme-linked immune-sorbent assay (cELISA) was used to test the samples. Prevalence (%) along with 95% confidence intervals (CI) was calculated, and a univariable analysis was conducted to screen different risk factors. Furthermore, a binary logistic regression model was built at the animal and flocklevel (Table 3). In total, 147 (28.8%, CI 24.9, 33.0) flocks tested positive for the CCPP and the seroprevalence ranged from 10.0 to 53.8% in various governorates (p = 0.001). The prevalence was significantly (P<0.001) high in goats (28%, CI 23.8, 32.5) as compared to sheep (13.1%, CI 24.9, 33.0). At flock level; the open herds (OR 2.08, 1.33, 3.27), having a location in the coastal regions (OR 1.70, 1.14, 2.53) and flock size of more than 100 animals (OR 2.45, 1.54, 3.91) were the significant risk factors for CCPP in Oman. At the animal level; goats (OR 2.87, CI 2.17, 3.81), and small ruminants above the age of one year (OR 2.23, CI 1.38-3.59) were found more likely to acquire CCPP. We suggest that a control program based on the changes in the management system to minimize the risk factors and a possible mass vaccination should be devised to check CCPP in Oman. [ABSTRACT FROM AUTHOR]

Brucellosis is a neglected endemic zoonosis in Bangladesh and has a significant impact on public health and animal welfare of dairy farming as well as dairy farm economics. A cross‐sectional study was conducted to evaluate the seroprevalence of and risk factors for brucellosis in dairy cattle in the Chittagong metropolitan area (CMA) of Chittagong, Bangladesh. We collected serum samples (n = 158) from six randomly selected dairy farms in the CMA between February and November, 2015. The Rose Bengal Plate Test (RBPT) and a competitive ELISA (cELISA) were used as the screening and confirmatory tests respectively. Farm level and animal level demographic and risk factor data were collected using a questionnaire. The risk factors were analysed using a multivariable logistic regression with random effects. The overall seroprevalences of antibodies against brucellosis in cattle were 21.5% (34/158) and 7.6% (12/158) based on parallel and serial interpretation of the two tests respectively. Our results revealed that 20.3% (32/158) samples were positive using the RBPT and 8.9% (14/158) were positive using the cELISA. The within‐herd seroprevalence ranged from 10% to 26.3% and 5 to 20.7% using the RBPT and cELISA tests respectively. The odds of seropositivity were significantly higher in lactating cows (OR: 2.59; 95% CI: 1.02–6.55), cows producing less than 2 litres of milk (OR: 29.6; 95% CI: 4.3–353.8), cow producing 2–12 litres of milk (OR: 4.8; 95% CI: 1.1–33.4) and cows with reproductive disorders (OR: 3.2; 95% CI: 1.2–10.1). About 7.6% (12/158) and 1.3% (2/158) of cattle were found to be infected with acute and chronic brucellosis respectively. Based on these results, we suggest that cows that have reproductive disorders and are producing little milk should be prioritized for brucellosis screening in CMA. The screening tests should be used to control brucellosis in cattle in order to protect animal welfare, human health and to minimize the economic losses. [ABSTRACT FROM AUTHOR]

(1) Background: Anaplasmosis is an infectious disease in camels caused by an obligate intracellular bacterium that is transmitted by ticks. (2) Methods: A cross-sectional study was conducted during 2020 to study the seroprevalence of Anaplasma spp. among Camelus dromedarius in three governorates in Egypt and assess the associated risk factors. Serum samples from 365 camels were examined by a competitive enzyme-linked immunosorbent assay (cELISA) test. (3) Results: Overall, the seroprevalence of anaplasmosis among camels was 18.6%. Multivariable logistic regression was performed, and it was discovered that tick infestation, application of acaricides, grooming practice and body condition were potential risk factors for Anaplasma spp. infection (odds ratio > 1) in dromedary camels. In contrast, the locality in which the camels lived and their age were not significant effects with regard to the occurrence of anaplasmosis. (4) Conclusions: The current findings suggest that improvement of protective measures to limit the effects of the identified risk factors can help to reduce the spread of anaplasmosis among camels in Egypt.

The present study was performed to evaluate and compare the Fluorescence Polarization Assay (FPA) as a confirmatory test for bovine brucellosis in the province of Carchi - Ecuador, comparing it with a immuno-enzymatic assay cELISA which is a test prescribed as confirmatory diagnosis in the National Bovine Brucellosis Control Program for Ecuador. The study was carried out in the province of Carchi, which is located in the north of Ecuador in blood serum samples (n = 1000) obtained from female bovines older than 2 years with unknown health status and vaccination, belonging to 94 Agricultural Production Units. The serum was analyzed by the Rose Bengal test (RB) and the positive results were confirmed through cELISA, and then compared with the FPA considering as cutoff ≥ 89.9 mP. Of the total samples analyzed with RB, 94 were positive. Of these positive samples 77 were confirmed for both cELISA and FPA. In addition, 11 samples that were positive for RB and cELISA were negative for FPA. Likewise, 6 samples positive for RB were classified as negative for both cELISA and FPA. The concordance analysis showed that there is a moderate concordance between the cELISA and the FPA with an index of 0.472 (95% CI 0.179 – 0.765) between the two tests.

West Nile virus (WNV) is a vector-borne virus maintained in nature by a bird-mosquito cycle. However, it can occasionally and accidentally infect horses and human beings, leading to sometimes severe or even fatal outcomes in these species. Therefore, the monitoring of its circulation and disease occurrence is of relevance. Unfortunately, it is underdiagnosed or not diagnosed in several African counties, including Namibia, where no data is currently available for horses. In this study, 98 horses in three different stables in the Windhoek city area were investigated. They were found to have a seroprevalence of approximately 7%. Positive reactions were seen at all three stables, suggesting a greater than expected prevalence of the virus. This is the first report of serological evidence for the presence of the virus in horses in Nambia. Even though clinical signs were not reported in any of the stables from which the sera were derived, the seroprevalence to the virus suggests that horses with high genetic and/or economic value could benefit from vaccination against WNV. Because of the zoonotic potential of the virus, these findings are also of significance to human health authorities. [ABSTRACT FROM AUTHOR]

• Monoclonal antibodies (mAbs) were raised against Besnoitia besnoiti tachyzoites. • The mAbs were used to establish specific competitive Besnoitia ELISAs (Bb-cELISAs). • Bb-cELISA1 revealed high analytic and diagnostic sensitivity and specificity. • The novel competitive ELISA can be used as a multi-species test. Serological cross-reactions represent a serious problem in some currently available tests to diagnose Besnoitia infections in many species including cattle, caribou and donkeys. False-positive results are due to the low positive-predictive value of these serological tests for besnoitiosis. These tests therefore have clear limitations if large herds are screened in areas with low prevalence, since increased numbers of false-positive reactions require confirmatory testing by alternative serological methods, e.g. immunoblotting, which are time-consuming and create extra costs. To overcome this problem, we aimed to develop a highly sensitive and specific competitive ELISA (cELISA) using a panel of 12 monoclonal antibodies raised against the tachyzoite stage of Besnoitia besnoiti. A cELISA set up with one of these antibodies (Bb-cELISA1) was screened with a large panel of B. besnoiti -positive bovine sera to estimate the diagnostic sensitivity of the test. Sera from herds with Neospora caninum - or Sarcocystis spp.-infected cattle were used to estimate its diagnostic specificity. Relative to a reference standard, which combined the results obtained in a previously established highly sensitive and specific ELISA, in the immunofluorescence antibody test and in B. besnoiti tachyzoite and bradyzoite immunoblots, the new Bb-cELISA1 revealed a diagnostic sensitivity of 99.2% (95% confidence interval: 97.1–99.9%) and a diagnostic specificity of 99.9% (95% confidence interval: 97.7–100%). This novel assay was tested on a variety of proven Besnoitia -positive sera from other species, including B. besnoiti -infected cats, rabbits or Besnoitia bennetti -infected donkeys or Besnoitia tarandi -infected caribou. The results obtained with the new Besnoitia -cELISA for these animal species also corresponded almost perfectly with those of the reference tests, which included immunoblot and immunofluorescence antibody tests. In conclusion, the novel Besnoitia -cELISA represents a valuable tool for the diagnosis and control of bovine besnoitiosis and for studies on the epidemiology of Besnoitia infections in a variety of host species, including naturally exposed wildlife and experimental hosts. [ABSTRACT FROM AUTHOR]

The study determined the positivity rate of Trypanosoma evansi and Neospora caninum antibodies in water buffaloes in the province of Nueva Ecija, Philippines using Polymerase Chain Reaction (PCR) for T. evansi and competitive Enzyme-linked Immunosorbent Assay (cELISA) for N. caninum antibodies. A total of 100 whole blood and 100 serum samples were collected to test for T. evansi and N. caninum, respectively. Rotat 1.2 VSG gene was target using PCR for T. evansi detection. Neospora caninum antibody detection was done from the serum samples using cELISA test kit. Results revealed that the positivity rate of T. evansi in Nueva Ecija was 11% (11/100). The positive animals identified were from the municipalities of Muñoz (4/16; 25%), Sta. Rosa (3/13; 23.08%) and Talugtug (4/16; 25%). The seropositive rate of Nueva Ecija for N. caninum. was 46% (46/100), seropositive animals were identified in Cabanatuan City, 57.14% (4/7); Science City of Muñoz, 43.14% (22/51); Sta. Rosa, 40% (4/10); Sto. Domingo, 50% (6/12); and Talugtug 50% (10/20). The seropositivity rate of N. caninum and the presence of T. evansi in Nueva Ecija may contribute to the cases of abortions in the province and further studies should be employed to confirm the association of these organisms to abortion cases on water buffaloes.. [ABSTRACT FROM AUTHOR]

Acute Anaplasma ovis infections can cause severe clinical symptoms and might lead to significant economic losses in small ruminant flocks. However, little information has been provided related the prevalence of anaplasmosis in Angora goats. The study was designed to investigate the prevalence of Anaplasma ovis serologically (cELISA) and microscopically (Giemsa stained blood smears) among Angora goats from Duhok districts of the northern part of Iraq. A total of 92 blood samples were randomly collected from three localities of Duhok city; Zakho, Batel and Sumil during the study period from April to October 2009. The infection rate of A. ovis was 38.04% by Giemsa stained blood smear and 66.3% by cELISA. The prevalence of A. ovis in female goats was higher than that in males, but statistically not significant difference (P>0.05) by using both methods. The prevalence was also significantly higher (P