Hemagglutination Inhibition Test Research Articles

Monoclonal antibodies to hemagglutinin of influenza A/H7N3 virus (Orthomyxoviridae: Alphainfluenzavirus: Influenza A virus)

Variants of influenza virus A/H7 have the same high pandemic potential as A/H5. However, the information about the antigenic structure of H7 hemagglutinin (НА) is considerably inferior in quantitative terms to similar data for H5 НА.The aims of the study were development and characterization of the monoclonal antibodies (MAbs) panel for HA subtype H7 of the influenza A virus. Viruses were accumulated in 10-day-old chicken embryos. Purification and concentration of the virus, determination of protein concentration, preparation of MAbs and ascitic fluids, hemagglutination and hemagglutination inhibition (HI) tests, assessment of antibodies' activity in indirect enzyme-linked immunosorbent assay (ELISA), as well as determination of MAbs isotypes and neutralization reaction (NR) were carried out by standard methods. The obtained MAbs to А/mallard/Netherlands/12/2000 (H7N3) strain were studied in HI test with a set of strains of different years of isolation belonging to different evolutionary groups. MAbs had a reduced reactivity compared to the immunogen-virus for all the studied strains. Cross-interaction of MAbs 9E11 and 9G12 in HI test with influenza A/H15 virus has been observed. Influenza A agent with H7 HA variant could serve as a potential cause of a future pandemic. Development of the MAbs panel for subtype H7 HA is an urgent task for both veterinary medicine and public health. The obtained MAbs can be used not only for epitope mapping of the H7 HA molecule (currently insufficiently studied) and as reagents for diagnostic assays, but also for determining common («universal») epitopes in HA of different strains of this subtype.

The effect of CpG-ODN (Toll-like receptor 21 agonist) as an adjuvant for Newcastle disease vaccination in broiler chicken

The present study was conducted to evaluate the immune-stimulating effect of CpG-ODN (Toll-like-receptor-agonist21) on broiler chickens vaccinated against Newcastle disease. The motifs were examined with different vaccination programes using different inoculation routes. 270 (one-day-old) - chicks were divided into 6 groups. Group one as a negative control sham inoculated with PBS; group two was injected intramuscularly with CpG-ODN; group three was vaccinated with live ND vaccine by eye-drop route; group four was vaccinated by eye-drop route and injected with CpG-ODN; group five was vaccinated with Newcastle disease inactivated vaccine and injected intramuscularly with CpG-ODN; group six was vaccinated by eye-drop route with live attenuated vaccine dissolved in CpG-ODN. Antibodies responses were monitored based on hemagglutination inhibition test and ELISA assay for the detection of IgG. The average body weight gain and feed conversion ratio were estimated to evaluate CpG-ODN growth-promoting effect. All vaccinated groups supplied with CpG-ODN recorded higher antibodies titer than the only vaccinated group. Group 4 which vaccinated and inoculated intra-muscularly with CpG-ODN recorded higher antibodies titer than other vaccinated groups. Antibodies titer was similar in group five and six without significant difference and both were lower than group 4. ELISA assay confirmed the significance between the adjuvanted groups at 21 and 35 days old. There were no significant differences (p≥0.05) observed between the vaccinated and adjuvated groups in either average body weight gain or in feed conversion rate. In conclusion, CpG-ODN proved its efficiency as an adjuvant when co-inoculated with Newcastle disease vaccines via different administration routes.

Immunogenicity of inactivated Equine Influenza (H3N8) virus vaccine with different adjuvents in equine

Equine influenza virus (EIV), is of the important commen and important respiratory infectious diseases of horses. Vaccination can reduce the the prevalence and severity of disease. In the current study, a trial to improve the strength and duration of immune response through using of montanide oil ISA 206, saponin and Mycobacterium pheli extract as adjuvants for an egg grown inactivated EIV [A/equine/Alexandria/1/08 (H3N8)]. Safety and potency of the prepared vaccine studied by inoculation into guinea pigs and horses then evaluated using Haemagglutination inhibition (HI) test for humeral immune response. Cellular immune response evaluated in horses inoculated with inactivated EIVvaccine adjuvanted with Mycobacterium pheli extract using lymphocyte blastogenesis. The vaccines were safe for g.pigs and horses which inducing HI antibodies titer that persist for 10 months post-inoculation in vaccinated horses. EI vaccine adjuvanted with montanide oil ISA 206 was more potent than the other vaccines adjuvanted either with saponin or M.pheli extract. EI vaccine adjuvanted with montanide oil ISA 206 also keeps at 4oC for over one year and at room temperature for 6 months without effect on its immunogenic characters.

Comparison of hemagglutination inhibition tests, immunoperoxidase monolayer assays, and serum neutralizing tests in detecting antibodies against blue eye disease in pigs

Blue eye disease (BED) of pigs was identified in the early 1980s in La Piedad, Michoacan, Mexico. The causal agent is Porcine orthorubulavirus (PRV), which affects pigs of all ages, producing nervous, respiratory, and reproductive disorders. BED is geographically endemic to the center of Mexico, where 75% of the country's swine industry is concentrated. Due to its adverse effects on the swine industry and the risk of dissemination to other countries, it is essential to have reliable diagnostic methods for BED. The objective of this study was to establish the optimal conditions for three serological tests, hemagglutination inhibition (HI), immunoperoxidase monolayer assay (IPMA), and serum neutralization (SN), and to compare their sensitivity, specificity, kappa coefficient, and predictive values. Twelve different HI protocols (9408 tests), one SN protocol and one IPMA protocol (784 tests, each) were evaluated. Forty-nine sera were analyzed, and thirty-seven sera showed true positive results, while twelve showed true negative results. The kappa coefficient was used to assess the variation in each test. The best HI protocol registered a sensitivity and specificity of 89 and 100%, respectively, the IPMA test showed values of 85 and 100%, and the SN test registered a sensitivity of 91% and a specificity of 96%. One of the disadvantages of the HI test is that when chicken red blood cells (RBCs) are used, elution occurs in a short incubation time, which would decrease the specificity. The use of bovine RBCs increases the specificity of the testy and makes it more stable, but it decreases the sensitivity. The results of HI and SN revealed the importance of eliminating the complement system of the serum and removing other inhibitors to avoid test nonspecificity. The IPMA test does not use an active virus; hence, it is considered safe and does not present any risk of disseminating PRV.

Comparison of hemagglutination inhibition and hemagglutinin pseudovirus neutralization titres in relation to protection against influenza in a mouse model.

The hemagglutination inhibition (HI) test has long been used as a standard measure of antibody response for inactivated influenza vaccines. However, the HI test has limitations, such as insensitivity when using some H3N2 virus strains and failure to detect neutralizing antibodies that target regions distant from the receptor binding site. We therefore examined a hemagglutinin pseudovirus neutralization (PVN) test as a possible supplement or alternative to the HI test. We evaluated the association of HI or PVN titres with protection against influenza infection in mice based on morbidity (where the illness was defined as 25% body weight loss). We assessed this relationship using dose-response models incorporating HI or PVN titres as a variable. The morbidity was correlated with the pre-exposure titres, and such a correlation was well described by a modified dose-response model. The mathematical modelling suggests that PVN titres consistently show a stronger association with invivo protection as compared to HI titres in mice. Given our findings, the PVN test warrants further investigation as a tool for evaluating antibody responses to influenza vaccines containing hemagglutinin. The resulting models may also be useful for analyzing human clinical data to identify potentially protective antibody titres against influenza illness.

Trials for preparation and evaluation of a combined inactivated reassorted H5N1 and Escherichia coli O157 vaccine in poultry.

Background and Aim:Avian influenza (AI), which is one of the major respiratory diseases of poultry, and Escherichia coli (E. coli) have caused major economic losses around the world, including in Egypt. Therefore, in this study, we aimed to produce a vaccine from E. coli O157 and AI H5N1 formulated with Montanide ISA70 for the protection of poultry against both diseases.Materials and Methods:We divided one hundred 3-week-old chicks into four groups: Group 1 was vaccinated with prepared inactivated AI H5N1formulated with Montanide ISA70, Group 2 was vaccinated with inactivated E. coli formulated with Montanide ISA70, Group 3 was vaccinated with combined E. coli and AI H5N1 formulated with Montanide ISA70, and Group 4 was an unvaccinated control group. We measured the immune response using the HI (hemagglutination inhibition) test, enzyme-linked immunosorbent assay (ELISA), and the challenge test.Results:We found the three vaccines to be safe and sterile during all periods of examination and observation. The HI test showed that Group 1 exhibited specific antibody titers of 2.3 log2, 4.3 log2, 7.5 log2, 7.8 log2, 8 log2, and 8.1 log2 from week 2 to week 7, respectively, post-vaccination. Group 3 exhibited antibody titers of 3.3 log2, 5.8 log2, 7.8 log2, 8 log2, 8.3 log2, and 8.3 log2 from week 2 to week 7, respectively, post-vaccination. The immune response in both groups reached a high titer at week 6. The combined inactivated E. coli and AI H5N1 vaccine generated a higher immune response than the inactivated AI H5N1 vaccine, and a significant difference exists between the two groups. For Groups 2 and 3, the ELISA antibody titer exhibited its lowest value, 1996.5 and 2036.7, respectively, at week 1 post-vaccination; whereas, both groups exhibited the highest titers, 2227.7 (for Group 2) and 2287.3 (for Group 3), in week 3 post-booster. The ELISA for the combined inactivated E. coli and AI H5N1 vaccine had a higher titer than did the inactivated E. coli vaccine, and a significant difference exists between the two groups. Moreover, the protection rate was higher in Group 3, with 100% for E. coli and 90% for the AI H5N1 vaccine.Conclusion:Our findings demonstrate that producing a combined vaccine using E. coli and AI H5N1 formulated with Montanide ISA70 is recommended for protection against both diseases.

ВЫЯВЛЕНИЕ УСТОЙЧИВОСТИ К АНТИБИОТИКАМ У ВОЗБУДИТЕЛЯ ГИСТОФИЛЕЗА КРУПНОГО РОГАТОГО СКОТА Histophilus somni

ВЫЯВЛЕНИЕ УСТОЙЧИВОСТИ К АНТИБИОТИКАМ У ВОЗБУДИТЕЛЯ ГИСТОФИЛЕЗА КРУПНОГО РОГАТОГО СКОТА Histophilus somni

PROPERTIES OF EXPERIMENTAL SAMPLES OF VACCINE AGAINST AVIAN INFECTIOUS CORYZA

PROPERTIES OF EXPERIMENTAL SAMPLES OF VACCINE AGAINST AVIAN INFECTIOUS CORYZA

Pathological and Molecular Characterization of Newcastle Disease Virus Isolated from Gallus gallus in Java, Indonesia

Background: Known as the vital avian disease in Indonesia, Newcastle disease (ND) has surged all over Indonesia which cause an immense drop in poultry industry. This investigation intended to examine the pathological and molecular characteristics of the latest Newcastle disease virus (NDV) derived from Gallus gallus in Java, Indonesia.Methods: The samples were inoculated on the embryonic chicken eggs (ECEs), then identified using the hemagglutination test (HA test), insulated isothermal PCR (iiPCR) and hemagglutination inhibition test (HI test). It conducted the reverse transcription-polymerase chain reaction (RT-PCR) using MSF1 and MSF2 primers and followed by nucleotide sequencing for the fusion protein gene. B-cells epitope prediction analysis was applied using the Immune Epitope Database and Analysis Resource (IEDB) and VaxiJen v2.0 to predict its immunogenicity. In addition, it also performed the histopathological examination by applying hematoxylin and eosin staining.Result: The results identified two viruses, NDV-1 and NDV-2, based on the fusion protein gene. Significantly, the cleavage site motif of NDV1 is 112GRQGRL117 (avirulent) and whereas for NDV2 is 112RRRKRF117 (virulent). Furthermore, it also predicted the “CKMGSRPSTKNPAP” peptide from NDV1 could be an immunogenic epitope candidate with a BepiPred linear epitope prediction score of 17.08. In summary, it suggests that NDV1 isolate can be used as a reference for vaccine design in Indonesia.

Relationship Between the Serotypes and Hemagglutinin Gene Sequences of Avibacterium paragallinarum.

Avibacterium paragallinarum has been subtyped into three serogroups (A, B, and C) and nine serovars (A-1, A-2, A-3, A-4, B-1, C-1, C-2, C-3, and C-4) according to the Page and Kume schemes. Both schemes use the hemagglutination inhibition test for serotyping. However, the relationship between the hemagglutinin gene (HMTp210) sequences and serotypes of A. paragallinarum is still unclear. This problem is partly due to the lack of information on the complete HMTp210 sequence from the formal reference strain of Page serogroup B (strain 0222 or Spross). In this study, we determined the complete HMTp210 sequence of strain Spross. The sequence of Spross and those of other HMTp210 sequences retrieved from GenBank were used to conduct phylogenetic analyses to investigate the relationship between the serotypes and HMTp210 sequences of A. paragallinarum. Four phylogenetic clusters, designated clusters A-1, A-2, B, and C, were identified. Clustering based on complete HMTp210 sequences correlates with serotyping based on hemagglutination inhibition tests. Serovar A-2 was found to contain a chimeric HMTp210 gene that might have resulted from recombination between serovar A-1 and serovar C-1. In addition, phylogenetic analysis based on partial sequences (approximately nucleotides 1-1200) of HMTp210 was sufficient to discriminate between serogroups A, B, and C. These findings could be valuable for developing a molecular method for serotyping of A. paragallinarum.

Efficacy of two vaccines against recent emergent antigenic variants of clade 2.3.2.1a highly pathogenic avian influenza viruses in Bangladesh

H5N1 highly pathogenic avian influenza viruses (HPAIVs) have caused outbreaks in poultry in Bangladesh since 2007. While clade 2.2.2 and 2.3.4.2 HPAIVs have not been detected since 2012, clade 2.3.2.1a viruses have caused continuous outbreaks since 2012 despite the use of vaccines. In this study, we evaluated the efficacy of two H5 vaccines licensed in Bangladesh, RE-6 inactivated vaccine, and a recombinant herpesvirus of turkeys vaccine with an H5 insert (rHVT-H5), for protection against recent field viruses in chickens. We selected three viruses for efficacy tests (A/chicken/Bangladesh/NRL-AI-3237/2017, A/crow/Bangladesh/NRL-AI-8471/2017 and A/chicken/Bangladesh/NRL-AI-8323/2017) from 36 H5 viruses isolated from Bangladesh between 2016 and 2018 by comparing the amino acid sequences at five antigenic sites (A-E) and analyzing hemagglutination inhibition (HI) titers with reference antisera. The RE-6 and rHVT-H5 vaccines both conferred 80–100% clinical protection (i.e. reduced morbidity and mortality) against the three challenge viruses with no significant differences in protection. In addition, both vaccines significantly decreased viral shedding from infected chickens as compared to challenge control chickens. Based on these metrics, the current licensed H5 vaccines protected chickens against the recent field viruses. However, the A/crow/Bangladesh/NRL-AI-8471/2017 virus exhibited antigenic divergence including: several unique amino acid changes in antigenic epitope sites A and B and was a serological outlier in cross HI tests as visualized on the antigenic map. The continuing emergence of such antigenic variants which could alter the dominant antigenicity of field viruses should be continuously monitored and vaccines should be updated if field efficacy declines.

Aktivitas Ekstrak Meniran (Phyllanthus niruri linn) Sebagai Immunostimulator pada Ayam yang Divaksin Penyakit Tetelo

Newcastle Disease or tetelo is one of main problem in poultry Industry in Indonesia. Prevention such as biosecurity control and routin vaccination program have been conducted to overcome this problem, but they have not given any great impact. Phyllanthus Niruri L. or meniran is well known as immunostimulatory. This research was aimed to reveal effect of Phyllanthus Niruri L. extract on chicken vaccinated with live vaccine LaSota. Administration of Phyllanthus Niruri L. extract was conducted on three different time which were 7 days before vaccination, 1 days after vaccination, and 3 days before and after vaccination. The amount of Phyllanthus Niruri L. extract administered were 2 ml, 2.5 ml, and 3 ml orally. Data of antibody titre were collected for 4 weeks after the treatment. It was obtained by measuring the antibody through Haemagglutination Inhibition test each week. According to the result Phyllanthus Niruri L. extract could increase the amount of antibody titre against Newcastle Disease. The amount of Phyllanthus Niruri L. extract given that capable to induced maximum of antibody titre was administered 1 days after the vaccination with amount 2.5 ml. It is suggested that Phyllanthus Niruri L. extract should be administered post vaccination to boost antibody synthesis.

Serosurvey of Avian Influenza Viruses (H5, H7, and H9) and Associated Risk Factors in Backyard Poultry Flocks of Lahore District, Pakistan.

Rural poultry constitutes 56% of the total poultry population in Pakistan; however, epidemiological information about avian influenza viruses (AIVs) in backyard poultry flocks is lacking. A cross-sectional survey of villages of Lahore district was conducted from July 2009 to August 2009 using two-stage cluster sampling and probability proportional to size (PPS) sampling to estimate seroprevalence and its associated risk factors. A random selection of 35 clusters from 308 villages of Lahore were considered, and from each cluster, six chickens aged >2 months were selected. A total of 210 serum samples were collected and examined by the hemagglutination inhibition (HI) test for specific antibodies against AIV subtypes H5, H7, and H9. Overall weighted seroprevalence for AIVs was 65.2% (95% CI: 55.6–74.8%), and for subtype H5, H7 & H9 was 6.9% (95% CI: 10.8–23.0%), 0% (95% CI: 0–1.7%), and 62.0% (95% CI: 52.2–71.8%) respectively. However, none of the samples were positive for H7. The average flock size was 17.3 birds, and the main purpose of keeping poultry was for eggs/meat (70.6%, 95% CI: 59.7–81.4). A majority of them were reared in a semi-caged system (83%, 95% CI: 74.5–91.3). Backyard birds were received from different sources, that is, purchased from the market or received as a gift from friends or any NGO, and were 5.7 times more likely to become avian influenza (AI) seropositive than those that were not exposed to these sources (CI 95%: 2.0–716.0). Backyard birds which were received from different sources, that is, purchased from the market or received from friends or any NGO, were 5.7 times more likely to become AI seropositive compared to those that were not (CI 95%: 2.5–18.7). To reduce the risk of AIV in Pakistan, continuous surveillance of backyard poultry would be needed.

In Vitro and In Vivo Characterization of a Pigeon Paramyxovirus Type 1 Isolated from Domestic Pigeons in Victoria, Australia 2011.

Significant mortalities of racing pigeons occurred in Australia in late 2011 associated with a pigeon paramyxovirus serotype 1 (PPMV-1) infection. The causative agent, designated APMV-1/pigeon/Australia/3/2011 (P/Aus/3/11), was isolated from diagnostic specimens in specific pathogen free (SPF) embryonated eggs and was identified by a Newcastle Disease virus (NDV)-specific RT-PCR and haemagglutination inhibition (HI) test using reference polyclonal antiserum specific for NDV. The P/Aus/3/11 strain was further classified as PPMV-1 using the HI test and monoclonal antibody 617/161 by HI and phylogenetic analysis of the fusion gene sequence. The isolate P/Aus/3/11 had a slow haemagglutin-elution rate and was inactivated within 45 min at 56 °C. Cross HI tests generated an R value of 0.25, indicating a significant antigenic difference between P/Aus/3/11 and NDV V4 isolates. The mean death time (MDT) of SPF eggs infected with the P/Aus/3/11 isolate was 89.2 hr, characteristic of a mesogenic pathotype, consistent with other PPMV-1 strains. The plaque size of the P/Aus/3/11 isolate on chicken embryo fibroblast (CEF) cells was smaller than those of mesogenic and velogenic NDV reference strains, indicating a lower virulence phenotype in vitro and challenge of six-week-old SPF chickens did not induce clinical signs. However, sequence analysis of the fusion protein cleavage site demonstrated an 112RRQKRF117 motif, which is typical of a velogenic NDV pathotype. Phylogenetic analysis indicated that the P/Aus/3/11 isolate belongs to a distinct subgenotype within class II genotype VI of avian paramyxovirus type 1. This is the first time this genotype has been detected in Australia causing disease in domestic pigeons and is the first time since 2002 that an NDV with potential for virulence has been detected in Australia.

Cross-reaction of seasonal influenza vaccine immune serum against Eurasian Avian-like H1N1 swine influenza virus

Objective: To evaluate the cross-reaction of seasonal influenza vaccine immune serum against Eurasian avian-like H1N1 swine influenza virus. Methods: Nine human infected Eurasian avian-like H1N1 swine influenza virus strains were obtained from national influenza surveillance network laboratories in Jiangsu, Hebei, Shandong, Yunnan, Hunan, Fujian and Tianjin provinces, and their genetic characteristics of hemagglutinin were analyzed by deep sequencing. 30 volunteers were recruited respectively from children (2-5 years old), adults (24-57 years old) and elderly (60-84 years old) who received 2019-2020 seasonal influenza vaccine in Anning city, Yunnan Province in October 2019, and serum samples were collected before and 1 month after vaccination. The hemagglutination inhibition test was used to evaluate the cross-reaction of serum before and after immunization against 4 strains of human infection with Eurasian avian-like H1N1 swine influenza virus isolated since 2015. Results: The homology of hemagglutinin genes of 9 Eurasian avian-like H1N1 swine influenza viruses was similar, but the difference of hemagglutinin heavy chain and light chain amino acid genes with A (H1N1) pdm09 (vaccine strain) were 90-101 and 24-30 amino acids respectively. The antibody titer of vaccine strain antiserum to vaccine strain was 2 560; the antibody titers of the vaccine strain antiserum to Eurasian avian-like H1N1 swine influenza virus and the Eurasian avian-like H1N1 swine influenza virus antiserum to vaccine strain were same as 640. The proportion of children, adults and elderly vaccinated with seasonal influenza vaccine with antibody titer ≥40 against vaccine strain was 90.0%, 70.0% and 73.3%, respectively; while the proportion merely were 46.7%, 36.7% and 33.3%-43.3% to 4 strains of Eurasian avian-like H1N1 swine influenza virus, respectively. Conclusion: Seasonal influenza vaccination does not provide effective cross-protection against Eurasian avian-like H1N1 swine influenza virus.

Spatial Distribution and Seroprevalence of Newcastle Disease in Kaduna State, Nigeria

Abstract Newcastle disease is one of the greatest constraints to the development of poultry production in Nigeria. In this study, the spatial distribution of Newcastle disease antibodies was determined using the Geographic Information System. A total of 400 serum samples were collected from chickens in districts around Kaduna Metropolis and screened for Newcastle disease virus antibodies using the haemagglutination inhibition test done according to the procedure of OIE (2002). The spatial distribution demonstrated that the highest antibody titre level for Newcastle disease was closely associated with communities that were at entry points (Zaria— Kaduna road; Nnamdi Azikiwe bypass road; Television garage; Abuja—Kaduna expressway and Kachia road) to the metropolis and houses that are closely situated near live bird markets signifying the importance of bird movements in the spread of the disease. About 31 % (124 of 400) of chickens had antibodies to Newcastle disease virus, with exotics breeds (32.0 %) with more Newcastle disease virus antibodies than local breeds (29.8 %). The Newcastle disease prevalence was 33.9 % (39 out of 115), 31.8 % (41 out of 129) and 28.2 % (44 out of 156) for Kaduna North, Kaduna South and Chikun Local Government Areas (LGA), respectively. A Newcastle disease prevalence of 29.8 % (54 out of 181) was recorded in the local chickens, while 32.0 % (70 out of 219) was obtained from the exotic chickens.

English

Avian influenza subtype H9 is epizootic in backyard poultry population of Pakistan. A cross-sectional study was conducted in two districts of Pakistan from two different agro-ecological zones selected as strata to estimate H9 seroprevalence, virus prevalence and to identify risk factors associated with H9 seroprevalence in backyard poultry. A stratified two-stage cluster sampling method was applied to collect samples (n=420) from two strata (agroecological zone). A total of 210 birds were selected as elementary units from 30 clusters (7x30) in each district from each stratum. Blood and oropharyngeal swab samples were collected from each bird. Sera samples were tested by Hemagglutination Inhibition Test (HI) to detect anti-H9 antibodies and swabs samples were tested by RT-PCR for H9. Overall seroprevalence of H9 in two strata was 57.88% (95% CI 34.88-80.87%), while virus prevalence was calculated to be 3.33%. Four factors were identified to be significantly associated (P<0.05) with H9 seroprevalence in the multivariable logistic regression analysis. The odds for H9 seropositivity were 9.43 times higher in flocks with fighting cockerel compared to those having no fighting cockerel (95% CI 4.68 -18.96). Existence of any pond, canal or any other water body near home premises also enhanced likelihood of H9 seroprevalence (OR: 10.04; 95% CI 3.27- 30.83). Backyard chicken raised with other bird species (like ducks, pigeons, or captive wild birds) had higher chances of H9 seropositivity than chicken raised alone (OR 4.12, 95% CI 1.35-12.56). Visit of any farm vehicle to the village had odds ratio of 20.96 (95% CI 5.74-76.51). Future surveillance is recommended to check the level of disease throughout the country

Discrepancy Between In-clinic and Haemagglutination-Inhibition Tests in Detecting Maternally-Derived Antibodies Against Canine Parvovirus in Puppies.

Canine parvovirus (CPV) is one of the most common causes of mortality in puppies worldwide. Protection against CPV infection is based on vaccination, but maternally-derived antibodies (MDA) can interfere with vaccination. The aim of this study was to evaluate the applicability of an in-clinic ELISA test to assess the CPV MDA in unvaccinated puppies and CPV antibodies in bitches, comparing the results with the gold standard haemagglutination inhibition (HI) test. Serum samples of 136 unvaccinated puppies were tested, along with sera of 16 vaccinated bitches. Five unvaccinated puppies were retested after vaccination. Both assays showed that the 16 vaccinated bitches had protective antibody levels against CPV. Conversely, significant discrepancies were observed for the MDA titers in unvaccinated puppies. Protective MDA titers were observed in 91.9% puppies using HI and in 40.4% by the in-clinic ELISA test, and only the latter one showed a decrease of MDA titers and percentages of protected puppies after the first weeks of age. Vaccination of five puppies with high HI and low in-clinic ELISA MDA titers resulted in seroconversion. Our results confirm the reliability of the in-clinic ELISA test in determining protective antibodies against CPV in adult dogs. Our findings also suggest that the in-clinic ELISA test kit may also be a useful tool to detect and quantify CPV MDA, thus allowing prediction of the best time to vaccinate puppies and reduction of the rate of vaccination failures due to interference by maternally-derived antibodies.

Sero-prevalence and serotypes of infectious bronchitis virus in free-range chicken in Plateau state, Nigeria

Globally, infectious bronchitis (IB) is an important respiratory viral disease responsible for enormous economic losses to poultry farmers. In Nigeria, limited reports on the prevalence and serotypes of the IB virus are available. Here, we investigated the prevalence and serotypes of infectious bronchitis virus (IBV) in chicken in Plateau State. A descriptive cross-sectional study was carried out involving 440 apparently healthy free-range local chickens sampled from eleven villages in four Local Government Areas (LGA) of Plateau State. Sera collected from the birds were screened for the presence of four IBV serotypes namely; Massachusetts (Mass), Arkansas (Ark), Connecticut (Con) and Delaware (De-072) using haemagglutinationinhibition (HI) test. In all, a prevalence of 82.95% (n = 365) was recorded. At LGAlevel, prevalence of 79.50%, 47.37%, 95.45% and 100% were recorded in Kanam, Mangu, Qua’an pan and Bassa LGAs, respectively. Based on serotype prevalence, Mass had 89.30% (n = 326); Ark 79.70% (n = 291); Con 88.20% (n = 322) while De-072 was 42.70% (n = 156). There were statistically significant associations between dominant serotype and the LGAs (p≤0.001). This study shows high prevalence of IBwith at least four strains of IBV present in free-range chicken flocks in Plateau State requiring attention for control measures.
 Keywords: Free-range chicken; Infectious bronchitis virus; Plateau state, Serosurvey, Serotype

Development and application of a novel triplex protein microarray method for rapid detection of antibodies against avian influenza virus, Newcastle disease virus, and avian infectious bronchitis virus.

Avian influenza virus (AIV), Newcastle disease virus (NDV), and avian infectious bronchitis virus (IBV) inflict immense damage on the global poultry industry annually. Serological diagnostic methods are fundamental for the effective control and prevention of outbreaks caused by these viruses. In this study, a novel triplex protein microarray assay was developed and validated for the rapid and simultaneous visualized detection of antibodies against AIV, NDV, and IBV in chicken sera. The AIV nuclear protein (NP), NDV phosphoprotein (P), and IBV nonstructural protein 5 (nsp5) were produced in a prokaryotic expression system, purified, and immobilized onto an initiator integrated poly(dimethylsiloxane) (iPDMS) film as probes to detect antibodies against these viruses in chicken sera. After optimization of the reaction conditions, no cross-reactivity was detected with infectious bursal disease virus, avian leukosis virus subgroup J and chicken anemia virus antisera. The lowest detectable antibody titers in this assay corresponded to hemagglutination inhibition (HI) titers of 24 and 21 for AIV and NDV, respectively, and to an IDEXX antibody titer of 103 for IBV, using the HI assay and IDEXX commercial ELISA kit as the reference methods. When156 serum samples were tested using the new assay, the HI test and the IBV IDEXX ELISA kit, the assay showed 96.8% (151/156), 97.4% (152/156) and 99.4% (155/156) diagnostic accuracy for detection of AIV, NDV and IBV antibody, respectively. The current study suggests that the newly developed triplex microarray is rapid, sensitive, and specific, providing a viable alternative assay for AIV, NDV, and IBV antibody screening in epidemiological investigations and vaccination evaluations.