In vivo antiviral effect of plant essential oils against avian infectious bronchitis virus.

BACKGROUND: Infectious bronchitis virus (IBV) leads to huge economic losses in the poultry industry worldwide. The high levels of mutations of IBV render vaccines partially protective. Therefore, it is urgent to explore an effective antiviral drug or agent. The present study aimed to investigate the in vivo anti-IBV activity of a mixture of plant essential oils (PEO) of cinnamaldehyde (CA) and glycerol monolaurate (GML), designated as Jin-Jing-Zi. RESULTS: The antiviral effects were evaluated by clinical signs, viral loads, immune organ indices, antibody levels, and cytokine levels. The infection rates in the PEO-M (middle dose) and PEO-H (high dose) groups were significantly lower than those in the prevention, positive drug, and PEO-L (low dose) groups. The cure rates in the PEO-M and PEO-H groups were significantly higher than those in the prevention, positive drug, and PEO-L groups, and the PEO-M group had the highest cure rate of 92.31%. The symptom scores and IBV mRNA expression levels were significantly reduced in the PEO-M group. PEO significantly improved the immune organ indices and IBV-specific antibody titers of infected chickens. The anti-inflammatory factor levels of IL-4 and IFN-γ in the PEO-M group maintained high concentrations for a long time. The IL-6 levels in the PEO-M group were lower than those in prevention, positive drug, and PEO-L groups. CONCLUSION: The PEO had remarkable inhibition against IBV and the PEO acts by inhibiting virus multiplication and promoting immune function, suggesting that the PEO has great potential as a novel anti-IBV agent for inhibiting IBV infection.

Decrease in fertilization and cleavage rates, but not in clinical outcomes for infertile men with AZF microdeletion of the Y chromosome.

This study aimed to explore whether the presence of a Y chromosome azoospermia factor (AZF) microdeletion confers any adverse effect on embryonic development and clinical outcomes after intracytoplasmic sperm injection (ICSI) treatment. Fifty-seven patients with AZF microdeletion were included in the present study and 114 oligozoospermia and azoospermia patients without AZF microdeletion were recruited as controls. Both AZF and control groups were further divided into subgroups based upon the methods of semen collection: the AZF-testicular sperm extraction subgroup (AZF-TESE, n = 14), the AZF-ejaculation subgroup (AZF-EJA, n = 43), the control-TESE subgroup (n = 28) and the control-EJA subgroup (n = 86). Clinical data were analyzed in the two groups and four subgroups respectively. A retrospective case-control study was performed. A significantly lower fertilization rate (69.27 versus 75.70%, P = 0.000) and cleavage rate (89.55 versus 94.39%, P = 0.000) was found in AZF group compared with the control group. Furthermore, in AZF-TESE subgroup, the fertilization rate (67.54 versus 74.25%, P = 0.037) and cleavage rate (88.96 versus 94.79%, P = 0.022) were significantly lower than in the control-TESE subgroup; similarly, the fertilization rate (69.85 versus 75.85%, P = 0.004) and cleavage rate (89.36 versus 94.26%, P = 0.002) in AZF-EJA subgroup were significantly lower than in the control-EJA subgroup; however, the fertilization rate and cleavage rate in AZF-TESE (control-TESE) subgroup was similar to that in the AZF-EJA (control-EJA) subgroup. The other clinical outcomes were comparable between four subgroups (P > 0.05). Therefore, sperm from patients with AZF microdeletion, obtained either by ejaculation or TESE, may have lower fertilization and cleavage rates, but seem to have comparable clinical outcomes to those from patients without AZF microdeletion.

Development of a colloidal gold immunochromatographic strip for rapid detection of avian coronavirus infectious bronchitis virus.

Avian infectious bronchitis virus (IBV) still causes serious economic losses in the poultry industry. Currently, there are multiple prevalent genotypes and serotypes of IBVs. It is imperative to develop a new diagnosis method that is fast, sensitive, specific, simple, and broad-spectrum. A monoclonal hybridoma cell, N2D5, against the IBV N protein was obtained after fusion of myeloma SP2/0 cells with spleen cells isolated from the immunized Balb/c mice. The N2D5 monoclonal antibody (mAb) and the previously prepared mouse polyclonal antibody against the IBV N protein were used to target IBV as a colloidal gold-mAb conjugate and a captured antibody, respectively, in order to develop an immunochromatographic strip. The optimal pH and minimum antibody concentration in the reaction system for colloidal gold-mAb N2D5 conjugation were pH 6.5 and 30 µg/mL, respectively. Common avian pathogens were tested to evaluate the specificity of the strip and no cross-reaction was observed. The sensitivity of the strip for detecting IBV was 10-1.4522 EID50/mL. The strip showed a broad-spectrum cross-reactive capacity for detecting IBV antigens, including multiple IBV genotypes in China and all of the seven serotypes of IBV that are currently prevalent in southern China. Additionally, the result can be observed within 2 min without any equipment. The throat and cloacal swab samples of chickens that were artificially infected with three IBV strains were tested using the developed strip and the qPCR method; the strip test demonstrated a high consistency in detecting IBV via qPCR gene detection. In conclusion, the immunochromatographic strip that was established is rapid, sensitive, specific, simple, practical, and broad-spectrum; additionally, it has the potential to serve as an on-site rapid detection method of IBV and can facilitate the surveillance and control of the disease, especially in resource-limited areas.

Complete genome sequence of an infectious bronchitis virus chimera between cocirculating heterotypic strains.

To date, multiple serotypes and genotypes of infectious bronchitis virus (IBV) have been isolated and identified. In order to provide more information on the viral evolution of IBVs, a new virulent strain named GX-NN09032, isolated from Guangxi, China, in 2009, was sequenced, and phylogenetic and recombination analyses were conducted. Furthermore, potential recombination events associated with GX-NN09032 were found in four IBV strains, including GX-YL5, DY07, CK/CH/SD09/005, TC07-2. The present study suggested that GX-NN09032 might contribute to the emergence of modern IBV variants through recombination.

Continuous evolution of avian infectious bronchitis virus resulting in different variants co-circulating in Southern China.

Sixty field strains of avian infectious bronchitis virus (IBV) were isolated from chicken flocks in different regions of Guangxi from 1985 to 2012. Phylogenetic analysis of S1 subunit glycoprotein genes revealed that field isolates from 2009-2011 mostly belonged to the LX4 type, while those from 1985-2008 belonged to the HN08 type, and a few others belonged to the 4/91 type, the TW type and the Mass type. In addition, it is noteworthy that no obvious regional differences were found among these 60 strains isolated from six regions in Guangxi, while there was a high degree of sequence identity among the isolates in the same period of time.

Serotype and genotype diversity of infectious bronchitis viruses isolated during 1985-2008 in Guangxi, China.

The genetic diversity of the hypervariable region I of S1 gene (HVR I) of infectious bronchitis (IB) vaccine strains H120, Ma5 and 4/91 was compared to that of 26 infectious bronchitis virus (IBV) strains isolated from the field in Guangxi province of China during the years 1985-2008, and the field isolates were classified into five major genotypes. Monovalent antisera against three vaccine strains and seven field isolates of different genotypes were prepared by immunizing rabbits with mineral oil adjuvant preparations containing viruses propagated in chicken embryos. Virus neutralization (VN) tests were performed in tracheal organ cultures (TOCs) using these 10 strains with the antisera, and a one-way VN test was then used to compare the relationship of 10 monovalent antisera to the other 19 field isolates. As a result, seven different serotypes were classified based on the results of VN tests with the 26 isolates plus the three vaccine strains. We found that different serotypes were prevalent during different time periods, that more new serotypes have been prevalent in more recent years, and the prevalence of the original dominant serotype has been in constant decline since 2004. In addition, the concordance rate of the 26 field isolates between the S1 genotypes and serotypes was 57.7%.

Identification of a novel avian coronavirus infectious bronchitis virus variant with three-nucleotide-deletion in nucleocapsid gene in China.

A novel avian infectious bronchitis virus (IBV) variant, designated as GX-NN160421, was isolated from vaccinated chicken in Guangxi, China, in 2016. Based on analysis of the S1 gene sequence, GX-NN160421 belonged to the New-type 1 (GVI-1) strain. More importantly, three consecutive nucleotides (AAC) deletions were found in the highly conserved structure gene N. The serotype of GX-NN160421 was different from those of the commonly used vaccine strains. The mortality of the GX-NN160421 strain was 3.33%, which contrasted with 50% mortality in the clinical case, but high levels of virus shedding lasted at least 21 days. In conclusion, the first novel IBV variant with three-nucleotide-deletion in the N gene was identified, and this unique variant is low virulent but with a long time of virus shedding, indicating the continuing evolution of IBV and emphasizing the importance of limiting exposure to novel IBV strains as well as extensive monitoring of new IBVs.

Construction and Immunogenicity Comparison of Three Virus-Like Particles Carrying Different Combinations of Structural Proteins of Avian Coronavirus Infectious Bronchitis Virus.

Infectious bronchitis virus (IBV) poses massive economic losses in the global poultry industry. Here, we firstly report the construction and immunogenicity comparison of virus-like particles (VLPs) carrying the S, M and E proteins (SME-VLPs); VLPs carrying the S and M proteins (SM-VLPs); and VLPs carrying the M and E proteins (ME-VLPs) from the dominant serotype representative strain GX-YL5 in China. The neutralizing antibody response induced by the SME-VLPs was similar to that induced by the inactivated oil vaccine (OEV) of GX-YL5, and higher than those induced by the SM-VLPs, ME-VLPs and commercial live vaccine H120. More importantly, the SME-VLPs elicited higher percentages of CD4+ and CD8+ T lymphocytes than the SM-VLPs, ME-VLPs and OEV of GX-YL5. Compared with the OEV of GX-YL5, higher levels of IL-4 and IFN-γ were also induced by the SME-VLPs. Moreover, the mucosal immune response (sIgA) induced by the SME-VLPs in the tear and oral swabs was comparable to that induced by the H120 vaccine and higher than that induced by the OEV of GX-YL5. In the challenge experiment, the SME-VLPs resulted in significantly lower viral RNA levels in the trachea and higher protection scores than the OEV of GX-YL5 and H120 vaccines, and induced comparable viral RNA levels in the kidneys, and tear and oral swabs to the OEV of GX-YL5. In summary, among the three VLPs, the SME-VLPs carrying the S, M and E proteins of IBV could stimulate the strongest humoral, cellular and mucosal immune responses and provide effective protection, indicating that it would be an attractive vaccine candidate for IB.

[Quality evaluation of 3 sperm counting chambers by computer-assisted sperm analysis system].

OBJECTIVE: To compare 3 common sperm counting chambers by the computer-assisted sperm analysis (CASA) system and evaluate their precision in analyzing sperm density and motility. METHODS: We used latex bead solution at (20 +/- 5) x 10(6)/ml as analogue semen samples and analyzed the samples with Makler, Leja and Microcell counting chambers, 30 times with each chamber. And the average (x +/- s) and the coefficient of variation of sperm density were calculated by the CASA system. Meanwhile 54 semen samples collected from the outpatients analyzed with the 3 sperm counting chambers by the CASA system for the rates of forward movement and motility of the sperm. RESULTS: The averages of sperm density obtained with Makler, Leja and Microcell chambers were (25.90 +/- 3.97) x 10(6)/ml, (18.74 +/- 1.62) x 10(6)/ml and (20.35 +/- 2.55) x 10(6)/ml, the coefficients of variation were 15.31%, 8.64% and 12.54%, the rates of sperm forward movement were (46.54 +/- 17.09)%, (30.65 +/- 14.88)% and (30.49 +/- 13.21)%, and the rates of sperm motility were (59.75 +/- 16.12)%, (46.76 +/- 14.11)%, (43.11 +/- 14.02)% respectively. There were significant differences in average sperm density among the 3 groups (P < 0.05). The rates of sperm forward movement and motility obtained with the Makler chamber were significantly higher than those achieved with the Leja chamber and Microcell chamber (P < 0.05), but there were no significant differences between the latter two (P > 0.05). CONCLUSION: The rates of sperm density obtained with the 3 sperm counting chambers differed significantly. In the analysis of sperm motility, a higher rate can be achieved with the coverslip-pressed chamber than the capillary-drawn chamber.

The emergence of the infection of subgroup J avian leucosis virus escalated the tumour incidence in commercial Yellow chickens in Southern China in recent years.

A total of 81 clinical cases of suspected tumours were submitted to our laboratory from Yellow chicken farms in southern China during the years 2010 through 2017. The tumour-like tissue samples were closely examined for common oncogenic avian viruses in cell culture and further analysed using polymerase chain reaction (PCR). During 2010-2012, Marek's disease virus (MDV) mono-infection was found to be the dominant cause of the tumour incidences (52.4%, 11/21) followed by co-infection of MDV+ALVs (19.1%, 4/21). Starting from the year 2013 the mono-infection of avian leucosis virus subgroup J (ALV-J) became the dominant agent of the tumour cases (83.3%, 5/6). During the most recent four years (2014-2017), co-infections involving ALV-J and MDV or between ALV subgroups have increased (23.4% and 18.5%, respectively), but each of the co-infections was still slightly lower than the ALV-J mono-infection incidence (33.3%). In contrast to the dominant MDV mono-infection cases before 2013, more recently, the emerging ALV-J mono-infection and ALV-J co-infections were largely responsible for the occurrence of avian virus-induced tumour incidences in the commercial local Yellow breeds of chickens in southern China. These results indicate that eradication measures against ALV on all chicken farms, especially on farms with the Yellow chickens, ought to be enhanced to reverse this trend.

Immune protection conferred by three commonly used commercial live attenuated vaccines against the prevalent local strains of avian infectious bronchitis virus in southern China.

Live attenuated vaccines are critical in the control of avian infectious bronchitis. It is necessary to know the protection conferred by commonly used commercial live vaccines. In this study, specific pathogen-free chicks were vaccinated with the commercial live vaccines H120, 4/91 and LDT3-A. Blood samples were collected at weekly intervals for the detection of IBV-specific antibodies and quantification of CD4+ and CD8+ T lymphocytes. At 21 days post-inoculation the vaccinated birds were challenged with the IBV prevalent local strains GX-YL5, GX-GL11079 and GX-NN09032, respectively. Trachea and kidney samples were collected at 5 days post-challenge for the detection of the virus. The results showed that the H120 group exhibited medium antibody levels, the lowest percentages of CD4+, CD8+ T lymphocytes and the highest viral loads. The 4/91 group showed the lowest antibody levels, but the highest percentages of CD4+, CD8+ T lymphocytes and the lowest viral loads. The LDT3-A group showed the highest antibody levels, the medium percentages of CD4+, CD8+ T lymphocytes and the medium viral loads. The protection rates of H120, 4/91 and LDT3-A groups were 41.7-58.3%, 75.0-83.7% and 66.7-75.0%, respectively. The present study demonstrated that the vaccines H120, 4/91 and LDT3-A could stimulate the immunized chicks to produce different levels of humoral and cellular immunity to resist the infection of IBV, but couldn't provide complete protection against the prevalent local strains of IBV in southern China. Also, the vaccine 4/91 offered the best immune protection among the three vaccines.

Protection against Virulent Infectious Bronchitis Virus Challenge Conferred by a Recombinant Baculovirus Co-Expressing S1 and N Proteins.

Avian infectious bronchitis virus (IBV) is the causative agent of infectious bronchitis, which results in considerable economic losses. It is imperative to develop safe and efficient candidate vaccines to control IBV infection. In the current study, recombinant baculoviruses co-expressing the S1 and N proteins and mono-expressing S1 or N proteins of the GX-YL5 strain of IBV were constructed and prepared into subunit vaccines rHBM-S1-N, rHBM-S1 and rHBM-N. The levels of immune protection of these subunit vaccines were evaluated by inoculating specific pathogen-free (SPF) chickens at 14 days of age, giving them a booster with the same dose 14 days later and challenging them with a virulent GX-YL5 strain of IBV 14 days post-booster (dpb). The commercial vaccine strain H120 was used as a control. The IBV-specific antibody levels, as well as the percentages of CD4+ and CD8+ T lymphocytes, were detected within 28 days post-vaccination (dpv). The morbidity, mortality and re-isolation of the virus from the tracheas and kidneys of challenged birds were evaluated at five days post-challenge (dpc). The results showed that the IBV-specific antibody levels and the percentages of CD4+ and CD8+ T lymphocytes were higher in the rHBM-S1-N vaccinated birds compared to birds vaccinated with the rHBM-S1 and rHBM-N vaccines. At 5 dpc, the mortality, morbidity and virus re-isolation rate of the birds vaccinated with the rHBM-S1-N vaccine were slightly higher than those vaccinated with the H120 control vaccine but were lower than those vaccinated with the rHBM-S1 and rHBM-N vaccines. The present study demonstrated that the protection of the recombinant baculovirus co-expressing S1 and N proteins was better than that of recombinant baculoviruses mono-expressing the S1 or N protein. Thus, the recombinant baculovirus co-expressing S1 and N proteins could serve as a potential IBV vaccine and this demonstrates that the bivalent subunit vaccine including the S1 and N proteins might be a strategy for the development of an IBV subunit vaccine.

Association of serum autoantibodies with pregnancy outcome of patients undergoing first IVF/ICSI treatment: A prospective cohort study.

The relevance of antiphospholipid (aPL), antinuclear (ANA) or antithyroid (ATA) antibodies in women undergoing in vitro fertilization (IVF) or intracytoplasmic sperm injection (ICSI) are controversial. The present study aims to investigate which autoantibodies are associated with the pregnancy outcome of patients undergoing first IVF/ICSI treatment. A total of 3763 IVF/ICSI patients were recruited from January to December 2015. Forty-five patients positive for aPL presenting adverse outcomes in their first cycle received low-dose aspirin treatment before the second transfer. Logistic regression analyses were performed to assess any association between autoantibodies and IVF/ICSI outcomes. The aCL-IgG was significantly associated with live birth rate (OR 0.58, 95% CI 0.36-0.96, p<0.05) and miscarriage rate (OR 2.04, 95% CI 1.23-3.40, p<0.01). The aCL-IgM was associated with miscarriage rate (OR 2.14, 95% CI 1.29-3.54, p<0.01). The aß2GPI-IgG was associated with implantation rate and clinical pregnancy rate (OR 0.61, 95% CI 0.24-0.96, p<0.05; OR 0.40, 95% CI 0.13-0.87, p<0.05, respectively). After the low-dose aspirin treatment, the live birth rate (37.0% vs. 19.1%, p<0.05) increased significantly in patients with positive for aPL. In contrary, the aß2GPI-IgM, ANA, anti-thyroglobulin (aTG) and anti-thyroperoxidase (aTPO) antibodies had no association with IVF/ICSI outcome. It is suggested that the presence of aCL-IgG, aCL-IgM and aß2GPI-IgG might exert a detrimental effect on IVF/ICSI outcomes. Low-dose aspirin treatment could be useful for patients positive for these antibodies. Therefore, it is suggested that these antibodies should be assessed prior to IVF/ICSI treatment.

[Big Y chromosome not significantly influences outcomes of in vitro fertilization and embryo transfer].

OBJECTIVE: To evaluate the influence of big Y chromosome on the outcomes of in vitro fertilization and embryo transfer. METHODS: Data of 127 cycles of IVF/ICSI-ET, performed in our Reproductive Medicine Center from March 2001 to June 2003 were analyzed. The patients were divided into two groups according to the length of chromosome: Group A, 56 cycles with big Y chromosome RESULTS: No significant difference was observed in the quality of embryos and in the and Group B, 71 cycles with normal karyotype. rates of fertilization, cleavage, clinical pregnancy, implantation, miscarriage, ectopic pregnancy, dead infant delivery, malformation, CONCLUSION: Big Y chromosome has no significant influence on the baby boy delivery and baby girl delivery between the two groups. development of embryos and the outcome of pregnancy.

[Genotypes and serotypes of avian infectious bronchitis viruses isolated during 2009-2011 in Guangxi, China].

In order to investigate the prevalence and track genetic and antigenic evolutions of infectious bronchitis virus (IBV) and their prevalence in Guangxi, China since 1985, gene amplification and sequencing and virus neutralization (VN) test on chicken embryo tracheal organ cultures were used in genotyping and serotyping of 28 IBV isolates during 2009-2011 in Guangxi. The results of N gene sequencing and comparison showed that the 28 isolates and reference strains were classified into three groups, and most isolates belonged to group Ill, while the isolates in 1985-2008 belonged to groups IV and II. The data of VN test indicated that the 28 isolates belonged to 6 serotypes; among them, 71. 4% belonged to serotypes 1, 2, and 3, and 11 (39.3%) shared the same serotype with the current vaccine strains. Given the data of our previous study, it is found that prevalent serotypes and their proportions varied in different areas of Guangxi and during different periods. These data lay a good foundation for developing an oil-emulsified inactivated polyvalent vaccine containing local dominant serotypes for the effective prevention and control of infectious bronchitis.

Genetic diversity of spike, 3a, 3b and e genes of infectious bronchitis viruses and emergence of new recombinants in Korea.

The nucleotide sequences of a region including S1, S2, 3a, 3b and E genes of twenty-seven infectious bronchitis virus (IBV) isolates in Korea between 1990-2011 were determined and phylogenetic and computational recombination analyses were conducted. The sizes of coding regions of some genes varied among IBV isolates due to deletion or insertion of nucleotides; the nucleotide similarities of S1, S2, 3a, 3b and E genes among the 27 isolates were 75.9%-100.0%, 85%-100.0%, 64.0%-100.0%, 60.4%-100.0% and 83.1%-100.0%, respectively. According to phylogenetic analysis of S1 gene, the 27 isolates were divided into five genotypes, Mass, Korean-I (K-I), QX-like, KM91-like and New cluster 1. The phylogenetic trees based on the S2, 3a, 3b, E genes and S1-S2-3a-3b-E (S1-E) region nucleotide sequences did not closely follow the clustering based on the S1 sequence. The New cluster 1 prevalent during 2009 and 2010 was not found in 2011 but QX-like viruses became prevalent in 2011. The recombination analysis revealed two new S gene recombinants, 11036 and 11052 which might have been derived from recombinations between the New cluster 1 and QX-like viruses and between the K-I and H120 (vaccine) viruses, respectively. In conclusion, multiple IBV genotypes have co-circulated; QX-like viruses have recurred and new recombinants have emerged in Korea. This has enriched molecular epidemiology information of IBV and is useful for the control of IB in Korea.

Molecular characterization of major structural protein genes of avian coronavirus infectious bronchitis virus isolates in southern china.

To gain comprehensive genetic information of circulating avian coronavirus infectious bronchitis virus (IBV) isolates in China, analysis of the phylogenetic tree, entropy of the amino acid sequences, and the positive selection as well as computational recombinations of S1, M and N genes of 23 IBV isolates was conducted in the present study. The phylogenetic trees based on the S1, M and N genes exhibited considerably different topology and the CK/CH/LSC/99I-type isolates were the predominant IBVs based on the phylogenetic analysis of S1 gene. Results of entropy of amino acid sequences revealed that the S1 gene had the largest variation; the M gene had less variation than the N gene. Positive selections were detected in not only S1 but also M and N gene proteins. In addition, five S1 gene recombinants between vaccine strain 4/91 and CK/CH/LSC/99I-type field isolate were confirmed. In conclusion, multiple IBV genotypes co-circulated; genetic diversity and positive selections existed in S1, M and N genes; 4/91 vaccine recombinants emerged in China. Our results show that field IBVs in China are continuing to evolve and vaccine strains may have an important role in the appearance of new IBV strains via recombination. In addition, the present study indicates that IBV evolution is driven by both generations of genetic diversity and selection.

Comparative genomics of Korean infectious bronchitis viruses (IBVs) and an animal model to evaluate pathogenicity of IBVs to the reproductive organs.

The K-I and nephropathogenic K-II genotypes of infectious bronchitis virus (IBV) have been isolated since 1995 and 1990, respectively, in Korea and commercial inactivated oil-emulsion vaccines containing KM91 (K-II type) and Massachusetts 41 strains have been used in the field. To date, genomic analyses of Korean IBV strains and animal models to test the pathogenicity of Korean IBVs to the reproductive organs have been rare. In the present study, comparative genomics of SNU8067 (K-I type) and KM91 IBVs was performed, and an animal model to test the pathogenicity of SNU8067 was established and applied to vaccine efficacy test. The genome sizes of SNU8067 (27,708 nt) and KM91 (27,626 nt) were slightly different and the nucleotide and amino acid identities of the S1 (79%, 77%), 3a (65%, 52%), and 3b (81%, 72%) genes were lower than those of other genes (94%-97%, 92%-98%). A recombination analysis revealed that SNU8067 was a recombinant virus with a KM91-like backbone except S1, 3a, and 3b genes which might be from an unknown virus. An SNU8067 infection inhibited formation of hierarchal ovarian follicles (80%) and oviduct maturation (50%) in the control group, whereas 70% of vaccinated chickens were protected from lesions.

[Evaluation of antigenic relationship of Guangxi isolates of infectious bronchitis virus].

Monovalent antisera of 3 vaccine strains and 7 representative field isolates were prepared based on the comparison of genetic diversity of the hypervariable region I of S1 gene (HVR I from 3 infectious bronchitis (IB) vaccine strains (H120, Ma5 and 4/91) ,one reference strain M41 and 26 IB field isolates. These 30 strains were classified in 7 different genotypes, respectively. Virus-neutralizing test on tracheal organ cultures (TOC) with chicken embryo were used to evaluate relatedness values of the antigenicity based on the antibody titer, to analyze the antigenic relationships between the isolates and vaccine strains, as well as to determine the serotypes of 26 IB viruses isolated from the field in Guangxi between 1985 and 2008. The results showed 30 strains were classified into 7 distinct serotypes and there were two predominant serotypes within the 26 isolates, serotypes 1 (totally 13 isolates) and serotype 2 (totally 5 isolates), respectively. In addition, there were some differences observed between the results of serotyping and the genotyping (including the S1, N, M and 3'UTR). The results of the study demonstrated that there were different predominant serotypes and multiple serotypes of IBV circulated in Guangxi in recent years, antigenic variation existed between Guangxi field isolates and vaccine strains.

[Genetic variation of S1 gene hypervariable region I of infectious bronchitis viruses isolated in different periods in Guangxi].

The S1 gene hypervariable region I (HVR I) of 22 infectious bronchitis virus (IBV) strains isolated in Guangxi during the period of 1985-2007 were sequenced and compared to that of the other IBV reference strains and the pigeon coronavirus isolates. A phylogenetic tree based on nucleotide sequences of HVR I of all the IBV showed that they were classified into 5 distinct Clusters. 16 out of 22 IBV isolates were grouped into Cluster I, and had higher homology with pigeon coronavirus isolates but lower homology with the Massachusetts (Mass) type vaccine strains. There were 4 and 3 amino-acid residues inserted at the sites of 33-34 and 34-35 respectively within HVR I in 15 isolates, except in isolate GX-NN6 there had 4 amino-acid residues inserted at the both sites; isolates GX-YL1 and GX-NN2 had close relationship with Mass type vaccine strains, and they shared Cluster II; isolates GX-G and GX-XD of Cluster III had close relationship with the Japanese strain JP Miyazaki 89 which was isolated at the same period; isolates GX-YL6 and GX-NN7 of Cluster V had close relationship with the European strain 4/91. The results showed that there were high phylogenetic diversity among the IBVs prevailed in the field in Guangxi resulting from the commonly occurred mutation or insertion within the S1 gene HVR I of the viruses, and majority of the isolates had lower homology with the commonly used Mass type vaccine strains. There was much higher homology among viruses isolated in the same period of time, but without distinct difference in geographical origins.