Horner syndrome-A rare complication after thyroidectomy for benign thyroid swelling.

Horner syndrome occurring after thyroidectomy is a rare entity and most of the reported cases have happened after surgeries on malignant thyroid swellings. In the present report, we describe a 27-year-old female who developed ptosis, miosis, enophthalmos, and anhidrosis on the second post-operative day after thyroidectomy for benign goiter. Post-operative ultrasound, computed tomography of neck, nerve conduction study, and electromyography of brachial plexus were unremarkable. Patient was kept on conservative management. She was given short course of Prednisolone orally for 2 weeks and was discharged on 150 mcg thyroxine. She had significant improvement in ptosis, miosis, and enophthalmos after six months. Horner syndrome is a rare but an important complication after thyroidectomy which may lead to cosmetic disfigurement. Surgeons should be well aware of this possibility and its presentation for timely recognition and management postoperatively. Early intervention should be done for any reversible cause, i.e., hematoma and oral steroids should be initiated as early as possible.

Femoral pseudoaneurysm in drug addicts--excision without revascularization is a viable option.

PURPOSE: To present a series of patients presenting with femoral pseudoaneurysm. RESULTS: Seventeen patients who presented with a femoral pseudoaneurysm during a 1 year period were included in this study. Parenteral drug abuse was the most common aetiological factor. The femoral artery was most commonly involved at its bifurcation. Sixteen patients (94%) had excision of the pseudoaneurysm with ligation of vessel and debridement without any revascularization and one patient (6%) had reverse saphenous grafting after excision and ligation of vessels. Four amputations (23%) were performed. Three (17%) were major limb amputations, which included one above knee and two below knee amputations. Four patients (23%) developed intermittent claudication. CONCLUSION: Excision of the pseudoaneurysm with ligation of vessels and wide debridement without immediate revascularization in infected pseudoaneurysms is a safe and effective treatment.

Dietary arginine intake alters avian leukocyte population distribution during infectious bronchitis challenge.

Although dietary arginine is a factor in immune function and disease resistance, the full range of effects has yet to be described. In this study, the effects of dietary arginine on leukocyte population changes were examined in the peripheral blood and the respiratory tract of chickens inoculated with infectious bronchitis virus (IBV) strain M41. At 2 wk of age, female line P2a White Leghorn-type chickens were randomly assigned to one of three diets with different arginine levels: a marginally deficient diet (0.5%), an adequate diet (1.0%), and a diet containing a high level of arginine (3.0%). All birds were inoculated with IBV at 4 wk of age, and then the peripheral blood and the respiratory lavage were collected at 1 and 7 d postinfection (DPI). The growth rate of birds that received 0.5% arginine was significantly lower than that of birds receiving 1.0 or 3.0% arginine, whereas the growth of the latter groups did not differ. The percentage and absolute number of heterophil (H) and the H/lymphocyte (L) ratio in the peripheral blood at 1 DPI significantly increased as dietary arginine increased. In the respiratory lavage at 1 DPI, the percentage of H also increased with dietary arginine increase. At 7 DPI, the percentage of CD8+ cells from birds fed the deficient diet was lower than those from birds fed the adequate diet and the diet containing a high level of arginine, whereas the cell surface density of CD8 antigen did not vary among groups. These results show that dietary arginine influences the character of the chicken cellular response to IBV and the distribution of responding leukocyte subpopulations in a target tissue for the infection.

Genetic and antigenic diversity in avian infectious bronchitis virus isolates of the 1940s.

In order to verify a commonly held assumption that only Massachusetts (Mass) serotype of infectious bronchitis virus (IBV) was prevalent in the United States between the 1930s (when IBV was first isolated) and the 1950s (when the use of commercial IBV vaccines began), we examined 40 IBV field isolates from the 1940s. Thirty-eight of those isolates were recognized as Mass serotype viruses based on their reactivity to Mass-specific monoclonal antibody (Mab) and neutralization by Mass-specific chicken serum. The remaining two isolates, N-M24 and N-M39, that did not react with Mass-specific Mab, resisted neutralization by Mass-specific chicken serum, and were neutralized only by homologous chicken antibody were identified as non-Mass IBV. When the first 900 nucleotides (nt) from the 5'-end of the spike (S1) glycoprotein gene and their deduced amino acid (aa) sequences were compared, the two non-Mass isolates differed from each other by 24% and 28%, respectively. In a similar comparison, the non-Mass viruses N-M24 and N-M39 differed from M28, a Mass-type isolate from the 1940s, by 21% and 22% (nt) and 28% and 27% (aa), respectively. These data indicate that antigenic and genetic diversity among IBV isolates existed even in the 1940s. Interestingly, when the N-terminal region of the S1 of M28 was compared to that of M41, a prototype Mass virus that has undergone countless number of in vivo and in vitro host passages, the two viruses differed by only 2% (nt) and 4% (aa). This finding suggests that frequent genetic changes are not inherent in all IBV genomes.

Maternal antibody to infectious bronchitis virus: its role in protection against infection and development of active immunity to vaccine.

Chicks hatched with high levels of maternal antibody had excellent protection (>95%) against infectious bronchitis virus (IBV) challenge at 1 day of age, but not at 7 days (<30%). This protection significantly (P<0.05) correlated with levels of local respiratory antibody and not with serum antibody.A high percentage of both maternal antibody-positive (Mab+) and maternal antibody-negative (Mab-) chicks failed to produce IBV antibody when vaccinated at 1 day of age by the intraocular route. In addition, Mab+ chickens had a weaker virus-neutralizing antibody response to a second IBV vaccination compared to Mab- birds (P<0.05). Mab+ chicks experienced a more rapid decline (P<0.01) in maternal antibody after 1-day-of-age vaccination compared to their unvaccinated counterparts.A monoclonal antibody-based blocking ELISA that measured antibody levels specific to S1 glycoprotein of IBV correlated well with virus-neutralizing antibody titers.

The exacerbating effect of infectious bronchitis virus infection on the infectious bursal disease virus-induced suppression of opsonization by Escherichia coil antibody in chickens.

Chickens infected with infectious bronchitis virus (IBV) and infectious bursal disease virus (IBDV) commonly develop secondary infection of the respiratory tract with Escherichia coli, resulting in significant economic losses. To understand the host factors that may contribute to the E. coli infection, we investigated macrophage-mediated E. coli phagocytosis, intracellular bacterial killing, and development of opsonizing antibody in previously uninfected chickens and in those infected with IBV, IBDV, and IBDV plus IBV. Macrophages from the peripheral blood and the respiratory tracts of chickens infected with IBV or IBDV plus IBV efficiently performed in vitro phagocytosis of E. coli in the presence of positive-control serum (i.e., E. coli antiserum produced in normal chickens). Those macrophages also had adequate bactericidal activity, indicating that IBV and IBDV infections had not affected their phagocytic activity or bactericidal function. The phagocytic activity of macrophages remained unaffected (P < 0.05) when the positive-control serum was replaced with E. coli antiserum produced in chickens infected with IBV alone. However, when E. coli antisera raised in IBDV-infected and, especially, that produced in IBDV plus IBV-infected chickens were supplemented, the percentage of phagocytosis and number of bacteria ingested per phagocyte were significantly (P < 0.05) less. These results indicate that although IBDV alone has the potential to markedly reduce opsonizing ability of antibody, this effect is significantly (P < 0.05) exacerbated by IBV infection.

Isolation and characterization of a novel antigenic subtype of infectious bronchitis virus serotype DE072.

Three infectious bronchitis virus (IBV) isolates, CU82792, CU82805, and CU82808, were recovered from sentinel chickens placed with three different layer flocks of a large commercial poultry farm in New York State. The three isolates were classified as members of the DE072 serotype on the basis of 1) their S1 genes could be amplified with only a modified primer designed for the DE072 serotype and 2) their restriction fragment length polymorphism patterns, after digestion with endonucleases HaeIII, BstyI and XcmI, were indistinguishable from that of DE072 virus. Additional characterization of one of the isolates, CU82805, revealed that its S1 gene bears approximately 96% identity at the nucleotide level and 94% identity at the amino acid level with DE072. Yet, in an in vitro reciprocal virus neutralization test, only a one-way neutralization was observed, i.e., antiserum to CU82805 neutralized DE072, whereas CU82805 was not neutralized by DE072 antiserum. Implications of these findings with regard to IBV diagnosis and immunization are discussed.

Systemic and local antibody responses to infectious bronchitis virus in chickens inoculated with infectious bursal disease virus and control chickens.

Serum and local (respiratory) antibody responses to infectious bronchitis virus (IBV) were studied in 5-wk-old white leghorn-type control chickens and chickens inoculated with infectious bursal disease virus (IBDV) at 1 day of age. Of the chickens inoculated with IBV alone, 93% had detectable levels of IBV antibodies in the sera and 87% had detectable antibodies in the respiratory lavage fluids. Compared to this group, only 73% and 65% of IBDV-IBV inoculated chickens had serum and respiratory antibodies, respectively. In chickens inoculated with IBV alone, the IBV antibodies were evenly associated with immunoglobulin classes IgM, IgG, and IgA, whereas the IBDV-IBV inoculated chickens mainly produced IgM-associated antibodies with low to negligible levels of IgA- and IgG-associated antibodies. These results suggest that the lack of adequate IgA- and IgG-associated antibody production in IBDV-infected chickens may account for their increased susceptibility to IBV infection.

Cytotoxic activity of cells recovered from the respiratory tracts of chickens inoculated with infectious bronchitis virus.

Previously uninoculated control chickens and chickens exposed to infectious bursal disease virus (IBDV) at 1 day of age were intranasally exposed to the M41 strain of infectious bronchitis virus (IBV) at 5 wk of age. Between 7 and 13 days after inoculation with IBV, cells were collected from the respiratory tracts of both groups of chickens and assayed for in vitro cytotoxic activity against a lymphoblastoid LSCC-RP9 target cell line using a 4-hr 51chromium-release assay (CRA). Compared to thymocytes from uninfected chickens, which were used as negative controls in the CRA, respiratory tract cells from both groups consistently displayed significant cytotoxic activity against LSCC-RP9 target cells. This cytotoxic activity, attributed to natural killer cells, was statistically more pronounced (P < 0.05) in IBDV plus IBV-infected chickens than in chickens inoculated with IBV alone.

Sequence analysis of gene 3, gene 4 and gene 5 of avian infectious bronchitis virus strain CU-T2.

We have previously reported the nucleotide sequences of gene 2 (spike (S) protein gene), gene 6 (nucleocapsid (N) protein gene), and the 3' end untranslated region of a novel avian infectious bronchitis virus (IBV) strain, CU-T2 [Jia et al. (1995) Arch. Virol. 140, 259 271]. In the present report we describe the sequences of the remaining genes of this strain (gene 3, 4 and 5) with the exception of gene 1 (RNA polymerase gene). Gene 3 contained three open reading frames (ORFs), 3a, 3b and 3c of 174, 195 and 282 nucleotides (nt), respectively. Gene 4 (membrane (M) protein gene) consisted of 749 nt with a single ORF of 687 nt. Gene 5 contained two ORFs, 5a and 5b, with 198 and 249 nt, respectively. Thus, in total, there were 7349 nt from the 5' end of S protein gene to the 3' end of the CU-T2 genome. The overall nt sequence homologies between gene 3, 4, and 5 of CU-T2 and those of other strains were between 84.1-90.8%, 85.8-88.8% and 90.4 96.4%, respectively. The predicted amino acid (aa) sequence homologies revealed that gene 3b and 5b were more conserved than 3a, 3c and 5a. Each individual gene of CU-T2 strain (with the exception of the RNA polymerase gene) had a different level of homology with the homologous gene of other strains, suggesting that the evolution of IBV strains in general has been a complex, and as yet, poorly understood process.

Characterization of a novel monoclonal antibody which identifies chicken secretory component.

A mouse monoclonal antibody (MAb) produced against chicken biliary IgA (bIgA) was characterized. This MAb, designated A63, reacted with purified chicken biliary IgA (bIgA) but not with serum IgA in ELISA. In Western blot analysis, it recognized an 80-kDa protein associated with bIgA. MAb A63, but not a chicken IgA-specific MAb, reacted strongly with chicken embryo allantoic fluid, a rich source of SC, in immunoblots. In immunohistostained sections of chicken intestines A63 stained intracytoplasmic vacuoles in the enterocytes consistent with goblet cells, whereas the IgA-specific MAb predominantly stained plasma cells in the lamina propria. Whereas the IgA-specific MAb only reacted with the homologous IgA, MAb A63 reacted with bile samples of chicken, turkey, and quail but not of duck or pheasant. These data collectively indicated that MAb A63 recognized an avian secretory component (sc).

Analysis of the serotype-specific epitopes of avian infectious bronchitis virus strains Ark99 and Mass41.

The Ark and Mass serotype-specific epitopes of infectious bronchitis virus were studied by immunofluorescence and immunoprecipitation of mutant and recombinant spike glycoproteins (S protein) expressed in mouse L cells. Serotype-specific monoclonal antibodies could bind to the recombinant proteins of Ark99 and Mass41 expressed from the chimeras in which the N-terminal thirds of the S1 sequences were reciprocally exchanged. Therefore, it appears that the respective serotype-specific epitopes of both strains were localized within the C-terminal two-thirds of the S1 proteins. Deletion and insertion of a five-amino-acid fragment on the S1 proteins of Ark99 and Mass41, altered the serotype-specific epitopes. This result implies that the five-amino-acid insertion on the S1 protein of the Ark serotype was involved in determining the conformation of the protein, probably acting as a spacer. In addition, it appears that an interaction between sequences of the N-terminal third and the remaining portion of the S1 protein determines the tertiary structure of the protein as well as the conformation of the serotype-specific epitope.

A novel variant of avian infectious bronchitis virus resulting from recombination among three different strains.

An antigenic variant of avian infectious bronchitis virus (IBV), a coronavirus, was isolated and characterized. This strain, CU-T2, possesses a number of unusual features, which have not been previously observed in IBV. The S1 glycoprotein of CU-T2 carries virus-neutralizing and serotype-specific epitopes of two IBV serotypes, Arkansas (Ark) and Massachusetts (Mass). Sequence analysis revealed that the virus, originally an Ark serotype, has acquired the Mass-specific epitope by mutation(s). This provides evidence that point mutations may lead to generation of IBV antigenic variants in the field. It was further observed that two independent recombination events involving three different IBV strains had occurred in the S2 glycoprotein gene and N protein gene of CU-T2, indicating that genomic RNA recombination in IBV may occur in multiple genes in nature. It was especially significant that a sequence of Holland 52 (a vaccine strain) had replaced half of the N gene of CU-T2. This proves that recombination among vaccine strains is contributing to the generation of IBV variants in the field. Based on these observations it is predicted that every IBV field isolate could have unique genetic nature. Therefore, several recently reported diagnostic and serotyping methods of IBV which are based on dot-blot hybridization, restriction fragment length polymorphism (RFLP), and polymerase chain reaction (PCR), may not reveal the true antigenic and/or genetic nature of IBV isolates, and may in fact yield misleading information.

Flow cytometric analysis of B cell and T cell subpopulations in specific-pathogen-free chickens infected with infectious bursal disease virus.

Lymphocytes obtained from the blood, spleen, and bursa of normal chickens and of chickens infected with infectious bursal disease virus (IBDV) were analyzed for phenotypic expression of CT4, CT8, and immunoglobulin cell surface markers. Single-cell suspensions were stained with monoclonal antibodies by an indirect immunofluorescent assay, and percent staining was quantitated by flow cytometry. Although an appreciable decline from control levels in the percentage of lymphocytes expressing IgM was detected in the spleen and bursa of infected chickens, the relative proportions of lymphocytes expressing CT4 and CT8 in peripheral blood and spleen remained unchanged following infection. These results suggest that whereas humoral immune depression by IBDV may be associated with lysis of antibody-producing B cells, cellular immune depression is not associated with a detectable change in the proportion of helper or cytotoxic/suppressor subpopulations of T lymphocytes.

A monoclonal antibody-based antigen capture enzyme-linked immunosorbent assay for identification of infectious bronchitis virus serotypes.

An antigen-capture enzyme-linked immunosorbent assay (C-ELISA) was developed for detection and identification of infectious bronchitis virus (IBV) serotypes Arkansas, Connecticut, and Massachusetts using monoclonal antibodies (MAbs) specific to the S1 glycoprotein of the respective serotype. The assay (designed as a double-antibody sandwich assay) gave the best results when the S1-specific MAb, antigen, and chicken serum were of the same serotype. However, when a group-specific (M glycoprotein-specific) MAb was used for antigen capture, a distinctive pattern of cross-reactivity was observed between the antigens and heterologous chicken sera, suggesting a complex distribution of epitopes on the IBV M glycoproteins. Treatment of antigen with NP40 enhanced the ELISA signal only when the M glycoprotein-specific MAb was used for antigen capture. Although C-ELISA was inconsistent in detecting IBV in chicken tissue homogenates, it was highly effective in detecting the virus in allantoic fluid after the homogenates were given one chicken embryo passage.

Oligonucleotide probes in infectious bronchitis virus diagnosis and strain identification.

Genomic RNA fingerprints of infectious bronchitis virus (IBV) strains M41 and Conn46 were prepared to identify T1 RNase-resistant oligonucleotides 'unique' to each of the two IBV strains. Such oligonucleotides were subsequently eluted from the gels and their nucleotide sequences determined. When oligonucleotide probes of those sequences were synthesized and used in a dot-blot hybridization assay, the probes lacked IBV strain-specificity and reacted with the RNAs of homologous as well as heterologous IBV strains. Based on these results, the methods used in this study need to be applied to a large number of oligonucleotide probes, to find one or a few that might be suitable as IBV strain- or serotype-specific oligonucleotide probes.

A monoclonal antibody blocking ELISA to detect serotype-specific infectious bronchitis virus antibodies.

A monoclonal antibody (Mab) blocking enzyme-linked immunosorbent assay (B-ELISA) was developed and compared to a conventional indirect ELISA (I-ELISA) and a virus-neutralization (VN) test for detection of specific antibodies to avian infectious bronchitis virus (IBV) serotypes. Sera used in this study were derived from chickens experimentally inoculated with the three most prevalent IBV serotypes, Arkansas (Ark), Connecticut (Conn), and Massachusetts (Mass). Overall, there was good correlation between the results of B-ELISA and the VN test. Both detected serotype-specific antibodies in chicken sera during primary and secondary phases of the immune response. Results of both tests indicated that the antibodies produced during the primary response to IBV serotypes are strongly serotype-specific. Those produced during the secondary response react more strongly with the homologous virus, but do exhibit some level of cross-reactivity with heterologous antigens. I-ELISA detected IBV group-specific and not serotype-specific antibodies. The B-ELISA which both offers the convenience of the conventional I-ELISA and the serotype specificity of the VN test, hold excellent promise for field application in IBV diagnosis and evaluation of response to IBV vaccines.

Production and characterization of monoclonal antibodies to three infectious bronchitis virus serotypes.

Three panels of monoclonal antibodies (MAbs) were prepared against the spike (S) proteins of infectious bronchitis virus (IBV) strains Arkansas 99, Connecticut 46, and Massachusetts 41. Based on enzyme-linked immunosorbent assay (ELISA), the MAbs were grouped into three categories: 1) group-specific, which reacted with a broad spectrum of homologous and heterologous IBV serotypes; 2) serotype-specific, which reacted only with strains of the homologous serotype; and 3) strain-specific, which reacted "selectively" with only certain strains of homologous and heterologous serotypes. MAbs that displayed serotype specificity were all specific to S1 fractions of the homologous serotype, confirming that epitopes that determine virus serotype are associated with the S1 protein. An excellent correlation was found when the results of IBV serotyping by MAb-based indirect ELISA were compared with those from the conventional virus-neutralization test. This confirms that the MAbs described here will serve as valuable tools in epizootiological studies and serotype-specific diagnosis of IBV infection.

Detection of viral antigen following exposure of one-day-old chickens to the Holland 52 strain of infectious bronchitis virus.

One-day-old specific-pathogen-free single comb White Leghorn chickens were inoculated by eyedrop with either 0.1 ml of phosphate buffered saline containing 10(4.3) EID50 of the Holland 52 strain of infectious bronchitis virus or normal allantoic fluid. Trachea, caecal tonsils and kidneys were removed from randomly selected birds at 0, 3, 5, 7 and 10 days post-inoculation (pi) and the presence or absence of viral antigen was detected utilizing virus isolation (VI), an indirect fluorescent antibody technique (IFA), or a streptavidin-biotin immunohistochemical (IH) technique. The presence of viral antigen as detected by the IH technique was also compared to histopathological changes in serial sections stained with haematoxylin and eosin. Detection of viral antigen occurred more frequently with VI than with IFA or IH. The IFA and IH techniques detected viral antigen with about the same frequency. Viral antigen was detected in the mucosa and submucosa of the trachea as early as 3 days pi, reached maximum levels at 5 days pi, and could still be detected in the mucosa at 10 days pi. In the kidney, viral antigen was not detectable by IFA or IH at 3 days pi, but could be visualized in distal convoluted tubules and collecting tubules at 5 days pi. At 7 days pi, antigen was detectable in the proximal convoluted tubules also. The presence of antigen in the caecal tonsils was sporadic, but it was detected in histiocytic cells and, occasionally, lymphoid cells of that organ.

A monoclonal antibody-based immunoperoxidase procedure for rapid detection of infectious bronchitis virus in infected tissues.

A monoclonal antibody (MAb)-based indirect immunoperoxidase (IP) procedure was applied to detect infectious bronchitis virus (IBV) antigen in frozen tissue sections. With this procedure, IBV antigen could be detected in chorioallantoic membranes (CAMs) within 15 hr postinoculation and in the respiratory tissues of chickens within 96 hr postinoculation. Endogenous peroxidases in chicken tissues were removed by treatment with periodic acid. The IP technique was highly sensitive, convenient, and economical. It allowed simultaneous evaluation of antigen-bearing cells and the general tissue morphology. The IP-stained slides also provided a permanent record.