Isolation of avian infectious bronchitis virus from experimentally infected chickens.

Virus was recovered from the faeces of chickens infected at three or four weeks of age for more than 20 weeks after infection with commercial vaccines or with the T strain of avian infectious bronchitis virus (IBV). Virus was not recovered from the trachea, liver, spleen, bursa or kidneys of T strain infected birds longer than 29 days after infection at which point IBV was recovered from the bursa of a single infected bird. In a subsequent experiment IBV was recovered from the caecal lymph nodes and faeces of one of five birds 14 weeks after infection with a commercial vaccine but no virus was isolated from the trachea, kidneys, duodenum, bursa, ovaries or testes of any of the five birds at this time.     

检测鸡传染性支气管炎病毒抗体的三种血清学方法的比较

应用酶联免疫吸附试验（ＥＬＩＳＡ）、琼脂扩散沉淀试验（ＡＧＰ）和鸡气管环培养中和试验（ＳＮｉｎＴＯＣｓ）三种常规血清学方法对实验鸡血样的鸡传染性支气管炎病毒抗体进行了检测。从实验鸡血样的检测结果表明，ＥＬＩＳＡ和气管环中和试验的灵敏度较好，而ＡＧＰ的灵敏度相对较差，但三者均有较好的特异性。对田间送检血样的检测结果表明，ＥＬＩＳＡ与气管环中和试验、ＥＬＩＳＡ与ＡＧＰ的一致性均较好，而气管环中和试验与ＡＧＰ的一致性较差。ＥＬＩＳＡ效价与气管环中和试验效价的相关系数为０．８４     

Establishment of persistent avian infectious bronchitis virus infection in antibody-free and antibody-positive chickens

Avian infectious bronchitis virus (IBV) causes a highly contagious and economically significant disease in chickens. Establishment of a carrier state in IBV infection and the potential for the persistent virus to undergo mutations and recombination in chicken tissues have important consequences for disease management. Nevertheless, whether chickens can maintain persistent IBV infection in the absence of reinfection from exogenous sources or the presence of antibody in the host can modulate virus persistence remains unclear. Indeed, whether or not IBV genome can undergo genetic changes during in vivo infection has not been demonstrated experimentally. In the present study, IBV shedding and tissue persistence were monitored in individual chickens maintained under strict isolation that precluded reinfection from exogenous sources. In the first of two experiments, intranasal exposure of 6-wk-old antibody-free chickens to IBV vaccine virus resulted in intermittent shedding of the virus from both trachea and cloaca of individual birds for up to 63 days. Also, the virus was recovered from the internal organs (spleen, gonad, kidney, lung, cecal tonsil, and cloacal bursa) of six of eight birds killed at various intervals between 27 and 163 days postinoculation (DPI). In the second experiment, IBV exposure of 1-day-old maternal antibody-positive chicks led to periodic virus shedding from the trachea and cloaca in all chickens until 77 days; however, internal organs (lungs and kidneys) of only one of seven birds (killed at 175 DPI) were virus positive, suggesting that presence of antibody at the time of infection protects internal organs from IBV infection. When the lung and kidney isolates of IBV from the latter experiment were compared with the parent-vaccine virus, no changes in their antigenicity, tissue tropism, or the nucleotide sequence of the S1 glycoprotein gene were observed. These findings indicate that, unlike the mammalian coronaviruses, propensity for frequent genetic change may not be inherent in the IBV genome.     

Resistance of chickens to challenge with the virulent Herts 33 strain of Newcastle disease virus induced by prior infection with serologically distinct avian paramyxoviruses.

Results indicate that some degree of protection from challenge by Newcastle disease virus (NDV)/Herts 33 was conferred on chickens by prior infection with PMV/turkey/Wisconsin/68, PMV/turkey/Ontario/6661/68, PMV/Netherlands/449/75 and PMV/parakeet/England/39/78 viruses, all of which are serologically related but distinguishable from NDV. Except for one bird which survived challenge three weeks after infection with Robin/Hiddensee/19/75, no protection was seen in chickens infected with other unrelated avian paramyxoviruses. In contrast to infection with NDV-B1, birds protected by infection with avian paramyxoviruses showed large increases in NDV haemagglutination inhibition (HI) titres after challenge. In these birds considerable increases in the homologous HI titres were also seen after challenge.     

The effect of route of infection and strain of virus on the pathology of Australian infectious bronchitis

The pathogenesis of infectious bronchitis in 4-week-old chickens was studied following administration of Australian T strain of IBV by the aerosol, conjunctival, intranasal, in-contact and drinking water routes.
Infected birds showed similar patterns of clinical signs, gross and histopathological lesions. While the birds infected by the aerosol route had earlier and slightly more severe respiratory tract disease it was concluded that the pathogenesis of infectious bronchitis virus nephritis was not significantly influenced by the route used to infect the birds.
In a second experiment, pathological changes produced in chickens infected with Australian S, A, A³ and T strains of IBV were compared in 18-day-old chickens infected by the aerosol route and housed in cold and warm environments. All 4 strains produced respiratory reactions and kidney lesions, and these changes were more severe and of longer duration in both respiratory tract and kidneys in the birds housed in a cold environment than those receiving supplementary heat.
The viruses differed in their virulence for the trachea and kidney and the effect of a virus on the respiratory trace was independent of the effect of that virus on the kidney.     

Serological examination and egg production of progeny of fowl experimentally infected with egg drop syndrome 1976 virus

Following EDS'76 virus (BC14 virus) infection of breeder chickens by the conjunctival route, vertical transmission occurred in the first week after infection. In the progeny which had been infected with EDS'76 virus by the vertical route, increasing haemagglutination inhibiting (HI) titres to BC14 virus and increasing numbers of birds with HI titres were observed from 3 weeks to 15 weeks of age. Sixty-one per cent of the hens and 77 per cent of the cocks had 2 log HI BC14 virus titres exceeding 4 at an age of 15 weeks. Some birds which han been serologically negative throughout the rearing period, seroconverted between 25 and 28 weeks of age. This phenomenon occurred in hens as well as in cocks. Simulation of stress twice during the laying period by injection of corticosteroid hormone did not increase the number of birds serologically positive to EDS'76 virus. EDS'76 was observed in the group of hens that was vertically infected, since egg production was significantly depressed between 28 and 34 weeks of age. Probably this was mainly the results of a production drop in the hens showing serconversion at 27 or 28 weeks of age. In this group of fowl vertically infected with EDs'76 virus, serologically positive birds appeared to be protected for the greater part to BC14 virus challenge at 50 weeks of age, while negative birds seemed to be fully susceptible. Chicks hatched from eggs collected in the third and fourth week after infection of the dams had maternal antibodies. Fertility and hatchability of apparently normally shelled eggs seemed not to be affected after BC14 virus infection of the dams. Intensive contact with contaminated faeces is probably an indispensable condition for lateral transmission of the virus.     

Comparison of duration of immunity in chickens infected with a live infectious bronchitis vaccine by three different routes.

Three groups of chicks were vaccinated by aerosol, intra-ocular and drinking water routes with a live infectious bronchitis (IB) vaccine. At one, two, six, 15 and 32 weeks after vaccination five birds from each group were sampled for testing for IB haemagglutination-inhibiting (HI) antibodies and challenged. Assessment of susceptibility to infection was measured by recovery of virus from individual tracheas and from kidney and gonad pools four days after challenge. Virus was isolated from all kidney and gonad pools of birds challenged one week after vaccination, the kidney and gonad pools of the drinking water vaccinates at two weeks, the kidney pool of the intra-ocular group at 15 weeks and all organ pools except the gonads of the intra-ocular group at 32 weeks. Tracheal resistance was found in most of the birds challenged one week after vaccination and in all the birds tested at two weeks but had begun to wane by six weeks after vaccination. No correlation was found between low HI antibody titres of individual birds and their susceptibility to challenge measured by reisolation of virus from the traches, but birds with titres over log2 6 were always resistant.     

Serum antibody responses of chickens following sequential inoculations with different infectious bronchitis virus serotypes

Sequential inoculations of chickens with different live infectious bronchitis virus (IBV) antigenic types had major effects on virus-neutralization (VN) and hemagglutination-inhibition (HI) serum antibody responses. Antibody production in IBV-inoculated chickens that were reinoculated 8 weeks later with heterologous virus was largely directed against the virus used for initial inoculation rather than the virus used for reinoculation. In addition, chickens inoculated sequentially with IBV produced a broadened spectrum of serum antibodies that reacted with IBV types to which the birds had never been exposed (JMK and Florida). Chickens inoculated sequentially with heterologous IBV tended to produce higher levels of cross-reacting antibody than birds given homologous virus inoculations. Levels of cross-reacting antibodies were lower than levels of specific antibodies directed against viruses that the birds had received. Limited studies indicated that birds with cross-reacting antibodies were not protected against challenge with the virus that the cross-reacting antibody was directed against. Implications of the research for interpreting serological data from commercial chicken flocks are discussed.     

Humoral antibody response and assessment of protection following primary vaccination of chicks with maternally derived antibody against avian infectious bronchitis virus

Commercially bred chicks with maternally derived antibody to avian infectious bronchitis virus (IBV) were hatched in isolated conditions and a number vaccinated within the first three weeks of life with live IBV strain H120. Humoral antibody responses were assayed by haemagglutination inhibition (HI) or neutralisation (SN) tests, and the degree of protection against challenge with the virulent Massachusetts M41 strain assessed on the basis of tracheal ciliary activity four days after challenge. Maternal antibody in unvaccinated chicks declined linearly with a mean half-life of five to six days based on both HI and SN tests; these chicks were protected against challenge until four weeks old. There was complete correlation between ciliary activity and histopathological findings, but little between protection and antibody titre. It was concluded that the optimum age for primary vaccination was about two weeks.     

SIMULATED NATURAL INFECTION OF CHICKENS WITH AUSTRALIAN LENTOGENIC NEWCASTLE DISEASE VIRUS AND SUBSEQUENT CHALLENGE WITH VIRULENT VIRUS

A total of 291 eight-week-old chickens were exposed to chickens infected with either of two Australian lentogenic strains (V4 and AVL NDV-1) of Newcastle disease virus (NDV). At 3 weeks after exposure, all chickens exposed to V4 infected chickens had developed haemagglutination-inhibition (HI) antibody. All chickens exposed to AVL NDV-1 virus infected chickens had developed HI antibody 5 weeks later. This sudden late appearance of HI antibody, to titres higher than those observed with V4 chickens, was explained by V4 virus being introduced to the AVL NDV-1 group of chickens. When groups of these chickens were challenged with Roakin virus (mesogenic NDV) at 3 weeks and Fontana 1083 virus (viscerotropic velogenic NDV) and Texas GB virus (neutrotropic NDV) at 3, 5, 10 and 21 weeks only three chickens developed clinical illness one of which died. These chickens were one AVL NDV-1 chicken contact challenged with Fontana 1083 virus at 3 weeks, one V4 chicken oronasally challenged with Texas GB virus at 5 weeks and one V4 chicken challenged oronasally with Fontana 1083 virus at 10 weeks. Susceptible non-vaccinated chickens died soon after challenge. Challenge by oronasal infection with 10(7.0) ELD50 of virus or contact with susceptible infected chickens enabled virulent virus to be isolated from most chickens and was accompanied by a large anamnestic increase in serum HI antibody.     

Serological comparison and antigenic relationships of seven serotypes of infectious bronchitis virus using the hemagglutination-inhibition test

The antigenic relationships, antigenic spectrum, and immunogenicity of seven isolates of infectious bronchitis virus (IBV) were examined using the hemagglutination-inhibition (HI) test. Because there was a discontinuity of antigenic relationships and a high degree of cross-reactivity among serotypes of IBV in cross-hemagglutination-inhibition tests, the range of antigenic spectrum used to group the serotypes with the HI test should be wider than the limits suggested by the plaque-reduction test. The HI test may provide valuable information in monitoring the immune status of a flock following vaccination when the area has a history of infectious bronchitis infection. It may also be used as a rapid diagnostic test if a flock is experiencing an outbreak of a disease caused by emergence of a new type of IBV. Interpretation of HI titers in evaluating immune status of chickens following infection with IBV depends on further cross-challenge and cross-protection studies of various types of IBV.     

Comparison of ciliary activity and virus recovery from tracheas of chickens and humoral immunity after inoculation with serotypes of avian infectious bronchitis virus

Cross-protection tests with homologous and heterologous serotypes of infectious bronchitis virus (IBV) were used to compare ciliary activity and virus recovery from tracheas of chickens. Validation of this technique included correlating the neutralization indices of antiserum obtained from some infected birds. Chickens were inoculated intratracheally with either the JMK or Connecticut (Conn) serotype of IBV. Three weeks later, infected and uninfected groups were challenged by the same route with homologous and heterologous virus. The JMK strain provided immunity against homologous challenge and the Conn strain, as indicated by good ciliary activity and lack of challenge virus recovery. The Conn strain provided only homologous protection, as ciliostasis occurred and virus was recovered after challenge with the JMK strain. In each case, antiserum to immunizing virus neutralized only the homologous virus. Controls were uniformly susceptible and lacked neutralizing antibody. A similar experiment with the Ark 99 serotype and a recent isolate (397) of IBV revealed complete cross-protection of the tracheas. Antiserum to each virus neutralized the homologous and heterologous virus in each case in reciprocal tests. The results indicate that these two viruses are closely related. The complete agreement between ciliary activity and virus isolation indicates that ciliary activity is a reliable, objective criterion upon which tracheal immunity can be judged in cross-protection tests.     

Duration of excretion of virulent Newcastle disease virus following challenge of chickens with different titres of serum antibody to the virus

Virulent Newcastle disease virus (NDV) was isolated from susceptible and immune chickens following intra-ocular challenge with the Essex '70 strain. Challenge virus was isolated from the trachea and cloaca of susceptible birds until they died 7 to 9 days after challenge. This virus was isolated from immunised chickens for up to 14 days after challenge. The duration of excretion was influenced by the prechallenge serum antibody titre to NDV. It persisted longest in chickens with titres of 2(3) to 2(7) and decreased in length and frequency from chickens with titres in the range 2(8) to 2(12). Chickens with pre-challenge titres of 2(3) to 2(5) developed 2- to 3- fold increases in post-challenge titres, whereas those with higher pre-challenge titres had smaller proportional increases in titre. Excretion of virulent virus from immunised birds should be considered in the development of Newcastle disease control programs.     

Protective antibody response produced by the chickens vaccinated with green coloured thermostable Newcastle disease virus

The efficacy of green-coloured (GC) I-2 Newcastle disease vaccine was determined in the present study. I-2 vaccine was mixed with a green coloured dye and stored at 4°C for 6 months while assayed for the virus infectivity at a monthly interval. Chickens were vaccinated with the GC vaccine by eye drop. Serum samples were collected from all birds before and after vaccination at weekly interval for 4 weeks and tested for haemagglutination-inhibition (HI) antibody against Newcastle disease virus (NDV). These chickens were challenged with NDV virulent strain four weeks after vaccination. The results showed that there was no difference between the infectivity titres of GC and uncoloured vaccines. However, chickens vaccinated with GC vaccine produced higher HI antibody titres than chickens vaccinated with uncoloured vaccine. Results from the challenge trial showed that all vaccinated chickens survived whereas all unvaccinated chickens died. The findings from this study have shown that the GC vaccine is safe and produced protective antibodies against NDV in vaccinated chickens. Wambura, P. N., 2008. Protective antibody response produced by the chickens vaccinated with green coloured thermostable Newcastle disease virus. Tropical Animal Health and Production.     

Investigations on the Influence of Helminth Parasites on Vaccination of Chickens against Newcastle Disease Virus under Village Conditions

Prevalence studies have shown that almost 100% of free-range chickens are infected with a wide range of parasites. The infections are mostly subclinical in nature, resulting in production losses and occasionally mortality. Newcastle disease (ND), on the other hand, results in high mortality rates during epidemics. ND is a limiting factor for increasing poultry production in many tropical countries, where frequent reports indicate vaccination failures. The aim of our study was to investigate the influence of helminths on the antibody response after vaccination against Newcastle disease of free-range chickens naturally infected with parasites. Sixty chickens were divided into six groups, of which three were vaccinated against ND with a live De Soto vaccine, while the other three remained non-vaccinated. One group within the vaccinated groups and the one within the non-vaccinated group was kept naturally infected with helminth parasites, while the other two groups in each set were dewormed with fenbendazole and niclosamide, and one of each of these groups was subsequently infected with Ascaridia galli. After vaccination, all the groups were followed for 5 weeks and their antibody titres were determined weekly using a HI test. All the birds were finally challenged 4 weeks after vaccination with a virulent velogenic ND virus obtained from a field outbreak. All the vaccinated chickens seroconverted and had high antibody levels after 3 weeks, but these dropped to low levels at 4 weeks after vaccination. After challenge, the antibody titres rose in the dewormed groups but not in the parasite-infected groups. After 5 weeks, all the parasite-infected animals had significantly lower antibody titres than the dewormed animals. All the vaccinated chickens survived the challenge infection, emphasizing the importance of the cellular immune response. Further studies are needed to examine the effects of the parasitic infection on protection against ND over a longer period.     

禽副粘病毒2型对SPF鸡免疫功能的影响

7日龄 SPF鸡经滴鼻、点眼感染 PMV- 2 ,可引起轻微的呼吸道症状 ,病理组织学观察可见气管黏液分泌亢进和轻微的淋巴细胞浸润 ;感染 PMV- 2后的 1天 ,2天 ,3天… 11天 ,定位采取气管、肺、肝、脾、肾、心、大脑、法氏囊、盲肠扁桃体等组织 ,检查病毒的分布规律 ,结果表明 ,PMV- 2在法氏囊、气管、肺、胸腺、脾、肾、大脑中均有分布 ;SPF雏鸡感染 PMV - 2后 ,接种鸡新城疫克隆 30疫苗 ,免疫后 5 - 30天 ,每 5天 1次检测血清中 ND的 HI抗体效价 ,结果表明 ,PMV- 2感染组比对照组的 HI抗体效价平均低 2 log2 ,统计结果显示差异极显著。雏鸡感染 PMV - 2后对新城疫疫苗的免疫应答有影响 ,从而可能抑制机体的免疫功能 ,危害养鸡生产。     

Immune response to oil-emulsion vaccines with single or mixed antigens of Newcastle disease, avian influenza, and infectious bronchitis

Inactivated Newcastle disease virus (NDV), avian influenza virus (AIV), and infectious bronchitis virus (IBV) antigens were evaluated for immunological efficacy in monovalent and polyvalent vaccines. Vaccinated broilers were bled for hemagglutination-inhibition (HI) tests at 1- or 2-week intervals. Half of the chickens were challenged with the Largo isolate of velogenic viscerotropic (VV) NDV at 8 weeks post-vaccination, and the remainder were challenged with the Massachusetts 41 strain IBV at 9 weeks post-vaccination. Newcastle disease HI titers were reduced significantly (P less than 0.05) from those of monovalent control vaccine groups when IBV antigen was emulsified in mixtures with low (1-3x) concentrated NDV or NDV and AIV antigens. Avian influenza HI titers were significantly (P less than 0.05) lower than those of the control monovalent groups when highly concentrated NDV was part of the polyvalent vaccine. Infectious bronchitis HI titers were higher than those of control monovalent groups in 13 of 15 vaccine groups when IBV antigen was in polyvalent formulations. VV NDV challenge killed all non-NDV vaccinates and induced increased HI titers in NDV vaccinates but no morbidity or mortality. Sixty of 80 IBV vaccinates experienced a fourfold or greater HI titer increase following challenge. All non-IBV vaccinates seroconverted at 1 week post-challenge.     

Study of protection by recombinant fowl poxvirus expressing C-terminal nucleocapsid protein of infectious bronchitis virus against challenge.

A stable recombinant fowl poxvirus (rFPV) expressing the C-terminal region (119 amino acids) of the nucleocapsid (N) protein of an infectious bronchitis virus (IBV) strain Ch3 was constructed by inserting the coding sequence within the thymidine kinase gene of fowl poxvirus (FPV) by homologous recombination. The N protein was expressed under control of the vaccinia virus promoter P7.5 in chicken embryo fibroblast cell cultures as seen in immunofluorescence assay and in rFPV-inoculated specific-pathogen-free (SPF) chickens by detecting antibodies with enzyme-linked immunosorbent assay (ELISA). A homologous IBV strain (Ch3) and two heterologous IBV strains (Ch5 and H4) were used to inoculate SPF chickens in a challenge to examine the protective efficacy of the rFPV. When the chickens were challenged with IBV Ch3 or Ch5, the control birds had respiratory signs of infections bronchitis, whereas all the vaccinated birds were clinically normal although low levels of the IBV infection were detected by a differential ELISA. In contrast, in the chickens challenged with IBV H4, all control birds and vaccinated birds suffered from the highly lethal IBV H4 infection. Our results suggest that the C-terminal 119 amino acid of the nucleocapsid expressed by FPV is a host-protective antigen and may induce cross-protective immunity against illness among some IBV strains.     

An enzyme-linked immunosorbent assay for the detection of antibody against avian influenza virus

An enzyme-linked immunosorbent assay (ELISA) was developed for detecting antibody to type A avian influenza (AI) virus. The sensitivity and group specificity of the AI-ELISA were compared with those of the agar-gel-precipitin test (AGPT) and the hemagglutination-inhibition (HI) test under conditions of both controlled and field exposure. During the course of temporal experimental infection (0-76 days) of specific-pathogen-free (SPF) chickens with AI subtype Hav9N2, the AI-ELISA was able to detect specific AI antibody as early as 8 days postinoculation (PI), and it measured rising levels of antibody through 35 days PI, at which time the chickens were re-exposed to AI virus. Conversely, AGP tests were negative through 35 days PI, and HI tests began to detect low levels of AI antibody only at 21 days PI. Following a secondary infection at 35 days PI with the same AI subtype, all tests measured rising levels of AI-specific antibody (35-76 days PI). However, the AGP test was positive at only the 7- and 14-day samplings postsecondary immunization. Under field conditions, the AI-ELISA was able to detect serum AI antibody in flocks from which highly pathogenic AI was isolated, but the AGP tests of these sera were negative.     

鸡滑液囊支原体活疫苗(MS-H株)同时免疫对新支二联活疫苗免疫效力影响的调查

为了解新支二联活疫苗和鸡滑液囊支原体活疫苗之间有无相互影响,选用70只1日龄SPF鸡随机分为4组进行比对试验,其中:G1组20只,联合免疫新支二联苗活疫苗和鸡滑液囊支原体活疫苗(MS-H株);G2组20只,为新支二联活疫苗单独免疫组;G3组20只,为传染性支气管炎病毒(IBV)攻毒组;G4组10只,为空白对照组.免疫后...