Sialodacryoadenitis virus-associated lesions in the lower respiratory tract of rats

Eight- to 10-week-old outbred Wistar rats were inoculated intranasally with 10(2.9) medium mouse lethal infective doses of sialodacryoadenitis (SDA) virus. Sham inoculated control rats and challenged rats were killed at 1 day intervals for the first 8 days, then on days 10, 12, 14, and 20. Typical lesions associated with SDA were seen microscopically in the salivary and lacrimal glands of inoculated rats. In addition, laryngitis, tracheitis, bronchitis, bronchiolitis, and multifocal alveolitis were present during the acute stages of the disease. Viral antigen was demonstrated in epithelial cells lining airways by immunofluorescence microscopy. SDA virus was recovered from the lower respiratory tract from days 2 to 6 post-inoculation (PI). Serum antibodies to SDA virus, but not to Sendai virus or Mycoplasma pulmonis were present in rats tested at day 20 PI. These findings demonstrate that during the acute stages of the disease, significant lesions do occur in the lower respiratory tract of SDA virus-infected rats.     

Sequential changes in the harderian and exorbital lacrimal glands in Wistar rats infected with sialodacryoadenitis virus

A sequential light and electron microscopic study of the exorbital and Harderian lacrimal glands was done on 2.5- to 15-month-old Wistar rats exposed to sialodacryoadenitis (SDA) virus. Typical coronaviral particles were readily demonstrated in cytoplasmic vesicles of Harderian and exorbital glands examined at 6 days post-inoculation. Lesions were seen in a relatively high percentage of lacrimal glands in infected animals of all ages, with no obvious age-related variations in the incidence and extent of changes. Lesions frequently persisted for a longer interval post-exposure in lacrimal glands than in salivary glands. The persistence of lesions commonly seen in Harderian glands was attributed, at least in part, to the cytotoxic effects of porphyrin-containing secretions released during the acute necrotizing stages of the disease. The persistence of lesions in some lacrimal glands indicates that they are useful tissues for microscopic examination for the retrospective provisional diagnosis of SDA. Persistent lesions also indicate that normal functions of these glands may be compromised for up to several weeks following outbreaks of SDA.     

Coronavirus infection in the laboratory rat: immunization trials using attenuated virus replicated in L-2 cells.

Sixty-nine specific pathogen-free male Wistar rats approximately eight weeks of age were used to evaluate the efficacy of an attentuated strain of sialodacryoadenitis (SDA) virus in providing protection against infection on subsequent challenge with virulent SDA virus. Fifty-four animals were inoculated intranasally with approximately 10(3.5) median cell culture infectious doses of the 25th passage of SDA virus in L-2 cells. Randomly-selected vaccinated animals were killed in order to evaluate the safety and efficacy of attenuated virus by histopathological examination of the salivary glands, lacrimal glands, and lower respiratory tract, and titration of sera for antibody to SDA virus. At three months and six months postvaccination (pv), animals were selected at random and challenged with virulent SDA virus. Seronegative, age-matched animals were also challenged, and served as controls. In animals examined at six to ten days pv, lesions were absent in submandibular and parotid salivary glands and lacrimal glands, but transient lesions were present in major airways of the lower respiratory tract. In a comparison of the incidence and extent of lesions, and antibody titers in challenged vaccinates and seronegative controls, lesions were minimal or absent in vaccinates compared to challenged naive rats, particularly in animals inoculated at three months pv. In addition, antibody titers in challenged vaccinates were much higher than were postinoculation titers in inoculated controls. In a comparison of lesions in salivary and lacrimal glands in vaccinated and control animals challenged at six months pv, there was a significant reduction in the number of animals without lesions in the vaccinated group (p = less than 0.0001).(ABSTRACT TRUNCATED AT 250 WORDS)     

Depletion of salivary gland epidermal growth factor by sialodacryoadenitis virus infection in the Wistar rat

Male and female Wistar rats 2 to 15 months of age were inoculated intranasally with sialoda-cryoadenitis (SDA) virus and killed at 8 to 21 days post-inoculation (PI). Submandibular glands were evaluated by light and electron microscopy, and levels of salivary gland epidermal growth factor (EGF) were quantitated by cytochemistry and competitive radioreceptor assay. Apical granules in the epithelial cells of the granular convoluted tubules (GCT) were selectively depleted during the acute and convalescent stages of the disease. In addition, levels of immunoreactive EGF were reduced in affected submandibular glands, especially at 8 to 14 days PI with SDA virus, but some evidence of EGF depletion was seen at up to 3 weeks PI. A corresponding transient depletion of EGF receptor reactive salivary EGF was seen between 1 and 3 weeks after experimental SDA infection. These studies suggest that a clinical (or subclinical) infection with SDA virus could have significant effects on experimental studies on EGF-dependent functions, including reproductive physiology and carcinogenesis.     

Coronavirus infections in the laboratory rat: degree of cross protection following immunization with a heterologous strain.

One hundred and twenty-one specific pathogen-free male Wistar rats eight to ten weeks of age were used to evaluate the efficacy of Parker's rat coronavirus (PRC) in affording cross protection on subsequent challenge with virulent sialodacryoadenitis (SDA) virus. Sixty-two animals were inoculated intranasally on day 0 and 21 days later with approximately 10(2) median tissue culture infective doses (TCID50) of the tenth passage of PRC replicated in L-2 cells. Animals were selected at random postvaccination to evaluate the safety and efficacy of PRC by histopathology, immunohistochemistry and serology. At three and six months postvaccination (PV), vaccinated and seronegative control rats were inoculated intranasally with approximately 10(3) TCID50 doses of virulent SDA virus. Challenged rats were then killed at 6, 10 and 14 days postchallenge and necropsied. Evaluations were based on lesion indices in lacrimal and salivary glands and respiratory tract, the presence of viral antigen by immunohistochemistry, and antibody response. Lesions were observed in rats killed PV, but in general, they were significantly reduced compared with those present in seronegative animals post-exposure to virulent SDA virus (p < or = 0.05). However, they were still considered to be an unacceptable level for a routine vaccination procedure. Potvaccination antibody titers to rat coronavirus were evident in all animals tested at three or six months prior to challenge with SDA virus.(ABSTRACT TRUNCATED AT 250 WORDS)     

Experimental sialodacryoadenitis virus infection in severe combined immunodeficient mice.

Mice with a severe combined immunodeficiency in B and T lymphocytes and natural killer cells (SCID-beige) were inoculated intranasally with sialodacryoadenitis (SDA) virus, a coronavirus of rats. Animals were killed at designated intervals and tissues were examined for evidence of viral infection by light microscopy and immunofluorescence microscopy. Based on these criteria, there was no evidence that these immunodeficient mice were susceptible to infection with SDA virus.     

A technique for inducing B-cell ablation in chickens by in ovo injection of cyclophosphamide.

The effect of cyclophosphamide (CY) treatment in ovo on avian B and T cells was studied. CY was injected in ovo on the 16th, 17th, and 18th days of incubation. Blood samples were collected periodically from CY-treated and nontreated birds after hatch and were used to measure blood lymphocyte responses to the T-cell and B-cell mitogens, concanavalin A and lipopolysaccharide (LPS), respectively. Additionally, flow cytometric analysis was used to determine the presence of B and T cells in peripheral blood, and birds were vaccinated with Newcastle disease virus (NDV) antigen at 3 wk of age and booster vaccinated at 5 wk of age. CY treatment reduced hatchability by 35%-40%, increased mortality by 3%-5% within the first 2 wk of life, and induced a significant retardation in body weight gains. At 2 wk of age, approximately 50% of CY-treated birds were devoid of B-cell mitogenic responsiveness while demonstrating significant T-cell mitogenic responsiveness. However, B-cell responses were observed at 4 and 6 wk from a small percentage of birds that were originally T-cell responsive and B-cell nonresponsive at 2 wk of age. Flow cytometric analysis of peripheral blood lymphocytes revealed that CY-treated birds had significantly less B cells (or were devoid of B cells) than the corresponding nontreated control birds. However, no significant difference in the T-cell percentage was observed between CY-treated and nontreated birds. CY-treated birds did not produce detectable antibodies specific for NDV during the first and second weeks postvaccination, as demonstrated by hemagglutination inhibition assay. However, antibodies were detected in some CY-treated birds 10 days postbooster. Those antibody-positive birds were found to be the same birds that had subsequently responded to the LPS mitogen on the blastogenesis microassay. This study indicates the importance of monitoring the B- and T-cell responses in CY-treated birds to identify those birds in which B-cell regeneration may have occurred.     

Effects of cyclophosphamide in newly hatched chickens after inoculation with avian nephritis virus

Effects of immunosuppression were compared in newly hatched chickens given cyclophosphamide (CY) after inoculation with avian nephritis virus (ANV). All CY-treated infected chickens died within 13 days after inoculation of the virus and had heavy urate deposits throughout the body. However, non-CY-treated infected, CY-treated noninfected, and non-CY-treated noninfected control chickens survived through the observation period. In a chronologic study, the value of serum uric acid in CY-treated infected chickens was more than 3 times higher than that in non-CY-treated infected chickens, and more than 9 times higher than in noninfected chickens. Serum uric acid values were coincident with the positive degree of ANV antigen in the tubular epithelial cells in the kidneys and with the severity of renal degeneration. Serologic and immunohistologic examinations did not reveal detectable antibody and IgG- and IgM-containing cells in the spleen and kidneys of CY-treated infected chickens. However, non-CY-treated infected chickens had an increased number of IgM- and IgG-containing cells and antibody against ANV on postinoculation day 6. These findings demonstrated that CY treatment enhanced the susceptibility of chickens to ANV infection.     

Experimental rabies in skunks: Immune response and salivary gland infection

Groups of striped skunks (Mephitis mephitis) were inoculated intramuscularly with graded doses of street rabies virus. At various intervals after inoculation, saliva and sera were tested for rabies virus and neutralizing antibodies, respectively. Skunks that developed rabies were killed in terminal stages of the disease and the following examinations were made: titers of virus and antibody in submandibular salivary glands and brain, extent of immunofluorescence in submandibular salivary glands, and histologic examination of various tissues.

Skunks that received inocula containing 4 × 10⁴ to 4 × 10⁵ mouse intracerebral lethal dose₅₀ (MICLD₅₀) had detectable serum neutralizing antibodies by 7–12 days postinoculation; however, most of the skunks that received lower doses (40 to 4 × 10³ MICLD₅₀) did not have detectable serum neutralizing antibodies until clinical signs began. In the salivary glands, slight and extensive immunofluorescence corresponded to high and low titers of tissue neutralizing antibody. Also low viral titers were associated with high tissue neutralizing antibody titers. There was a close correlation between viral titers in right and left submandibular salivary glands.

The results suggest that the immune response can impede the process of infection of the salivary glands resulting in lack of antigen or low amounts of antigen in this tissue. This could occur through interference with centrifugal neural transport of virus and/or neutralization of virus during transfer from neural elements to epithelial cells. Lack of infectious virus or low viral titers in salivary glands containing antigen and high levels of tissue neutralizing antibodies can be caused partly by postmortem virus neutralization (during viral titration).     

A pathological study of the salivary glands of rabid dogs in the Philippines

Rabies is a zoonotic disease caused by the rabies virus. While the salivary glands are important as exit and propagation sites for the rabies virus, the mechanisms of rabies excretion remain unclear. Here, we investigated the histopathology of the salivary glands of rabid dogs and analyzed the mechanism of excretion into the oral cavity. Mandibular and parotid glands of 22 rabid dogs and three control dogs were used. Mild to moderate non-suppurative sialadenitis was observed in the mandibular glands of 19 of the 22 dogs, characterized by loss of acinar epithelium and infiltration by lymphoplasmacytic cells. Viral antigens were detected in the mucous acinar epithelium, ganglion neurons and myoepithelium. Acinar epithelium and lymphocytes were positive for anti-caspase-3 antibodies and TUNEL staining. In contrast, no notable findings were observed in the ductal epithelial cells and serous demilune. In the parotid gland, the acinar cells, myoepithelium and ductal epithelium all tested negative. These findings confirmed the path through which the rabies virus descends along the facial nerve after proliferation in the brain to reach the ganglion neurons of the mandibular gland, subsequently traveling to the acinar epithelium via the salivary gland myoepithelium. Furthermore, the observation that nerve endings passing through the myoepithelium were absent from the ductal system suggested that viral proliferation and cytotoxicity could not occur there, ensuring that secretions containing the virus are efficiently excreted into the oral cavity.     

Pathogenesis of sialodacryoadenitis in gnotobiotic rats.

The pathogenesis of sialodacryoadenitis was studied in gnotobiotic CD rats inoculated intranasally with the causal virus. Virus replication was detected sequentially in the nasopharynx, tracheobronchial tree, cervical lymph nodes, submaxillary and parotid salivary glands, exorbital gland, and Harderian gland. Acute rhinitis appeared within 2 days after inoculation, and salivary glands had lesions in 4 days. Early changes in salivary and exorbital glands were characterized by necrosis of ductal epithelium, which rapidly progressed to widespread acinar necrosis, marked inflammation, edema and total effacement of glandular architecture. Harderian glands also had massive necrosis of tubuloalveolar units. Repair in all glands was characterized by marked squamous metaplasia of tubuloalveolar units. Repair in all glands was characterized by marked squamous metaplasia of ducts. Neutralizing and complement-fixing antibodies were detected in 7 days, and there was a concomitant decrease in tissue-virus titers. There was no detectable evidence for hematogenous spread of virus or for retrograde infection by way of major salivary ducts.     

Experimental rabies in skunks: effects of immunosuppression induced by cyclophosphamide.

Striped skunks (Mephitis mephitis) were inoculated with street rabies virus and immunosuppressed with several doses of cyclophosphamide. Control skunks were inoculated with street virus only. The skunks were killed in terminal stages of the disease and several tissues were collected for examination by immunofluorescence, light microscopy and viral titration. Sera collected at euthanasia from most of the principals did not contain detectable rabies neutralizing antibodies, whereas high titers occurred terminally in controls. Immunofluorescence was much more entensive in submandibular salivary glands of cyclophosphamide-treated than control skunks. Similarly, virus was isolated from this tissue more consistently and at higher titer from principals than from controls. Immunofluorescence was extensive in brains of all skunks (both groups), but virus was isolated consistently only from brains of cyclophosphamide-treated skunks. Most of the cyclophosphamide-treated skunks had very few inflammatory cells in brain and cerebrospinal ganglia. Neuronal degeneration occurred in dorsal root ganglia of both principals and controls. The results suggest that the immune response has no effect on the development of rabies-induced aggressive behavior, that the immune response may inhibit salivary gland infection and that it is not essential for the development of neuronal degeneration in dorsal root ganglia.     

Immunoperoxidase study of Aujeszky's disease in pigs

Viral antigen was detected by an immunoperoxidase technique in histological sections from pigs with Aujeszky's disease. The antigen was found mainly in association with focal necrosis in the cerebellum, tonsils, oral and nasal mucosa, salivary glands, lungs, liver, kidneys, pancreas, spleen and adrenal glands. Cells at the margin of the necrotic foci especially were strongly positive. Viral antigen was also demonstrated in the cerebral cortex and in the brain stem. Two types of intranuclear inclusion bodies were found to contain viral antigen and one type also contained viral nucleic acids. Inflammatory cells usually contained no viral antigen. The possible significance of some of these infected tissues in the excretion of the virus is discussed.     

Vaccination of cyclophosphamide-treated chickens against Newcastle disease virus infection

Newly-hatched chickens were treated with 3 mg of cyclophosphamide (CY) per day for 4 consecutive days. At 2 weeks of age, these chickens, together with a group of untreated controls, were vaccinated intranasally or subcutaneously with the La Sota strain of Newcastle disease virus (NDV). All chickens were challenged intranasally with the GB strain of NDV 2 weeks later. CY-treated, intranasally vaccinated chickens were highly resistant to NDV challenge, yet none of the chickens produced any detectable humoral antibodies to NDV; antibodies to NDV were detectable in the tracheal washings, however.     

Foot-and-mouth disease virus (FMDV) experimental infection: susceptibility and immune response of adult mice

Adult mice are susceptible to foot-and-mouth disease virus (FMDV) infection only under some experimental conditions. This paper report the results of pathogenesis studies on 4 different strains of mice (CF1, C3H, NIH-nude, BALB-c/J) infected with the cloned and uncloned 0(1)C strain of FMDV. High virus titers were detected in blood and pancreas 12-24 h after infection (p.i.); these persisted for up to 48 h p.i. in CF1 and BALB-c/J mice and 72 h p.i. in the two other mouse strains. Virus titers observed in other organs were lower than those found in blood. In pancreas, and occasionally in salivary glands, oropharynx, heart and testicles, viral antigen was detected by direct immunofluorescent assay. Circulating neutralizing antibodies appeared in CF1 and C3H mice at 72 and 96 h p.i. respectively, and their titers remained unchanged during the 30-day experimental period. Antibodies against viral infection-associated antigen (VIA) were detected for a shorter period. In animals irradiated with 1 LD 50 (total body irradiation), viremia persisted up to 14 days p.i. and a low antibody response was observed which began at the end of viremia. No differences in the response of mice to cloned or uncloned FMDV were observed.     

吸血蓖子硬蜱成虫唾腺莱姆病螺旋体重度感染的发现

为了解莱姆病螺旋体在蜱体内的发育生物学，采用直接免疫荧光试验、银染色法组织学及电子显微镜观察螺旋体，结果显示，感染莱姆病螺旋体的饥饿蓖子硬蜱雌虫未吸血与吸血0．5和1天唾腺中无螺旋体。在吸血开始后2天出现唾腺的轻度感染，3天出现唾腺的中度感染，4天和5天出现重度感染。这些结果表明，莱姆病螺旋体媒介蜱唾腺的感染程度随吸血时间的延长而迅速增高，并可在缓慢吸血期的中后期形成重度感染。莱姆病螺旋体在吸血期媒介唾腺内的大量存在，特别是重度感染的发现，为该病原螺旋体由蟀向脊椎动物宿主传播的唾液分泌途径假设提供了有力证据。本研究对莱姆病螺旋体在吸血蟀唾腺内分裂增殖的可能性进行了讨论。     

Exacerbation of murine respiratory mycoplasmosis by sialodacryoadenitis virus infection in gnotobiotic F344 rats

To test the hypothesis that sialodacryoadenitis virus infection could exacerbate respiratory mycoplasmosis in rats, four groups of 40 7- to 9-week-old gnotobiotic F344/N rats were given two intranasal inoculations 7 days apart: Mycoplasma pulmonis, then sialodacryoadenitis virus; M. pulmonis followed by sterile culture medium; medium initially, then virus; or two doses of medium. Immediately and 3, 5, 10, and 20 days after the second inoculation, the nasal passages, middle ears, larynges, tracheas, lungs, and salivary and lacrimal glands of four rats from each group were prepared for histologic examination, and the respiratory organs from four other rats were collected for quantitative culture of M. pulmonis and sialodacryoadenitis virus. To test statistically the effect of virus infection on mycoplasmosis lesions, we determined indices of the severity of respiratory tract lesions by subjective scoring. In rats given both organisms, indices of nasal and tracheal lesions were significantly (P less than 0.05) greater at 3 days and after than in rats given M. pulmonis alone, and middle ear, laryngeal, and lung lesion indices were significantly greater at 5 days and after. Rats given both mycoplasma and virus had significantly more mycoplasmal colony-forming units in the nasal passages at 3 days and after, and in the larynges, tracheas, and lungs at 10 and 20 days, than rats given only mycoplasma. These results show that sialodacryoadenitis virus infection can exacerbate respiratory mycoplasmosis in rats under experimental conditions; therefore, the virus probably also contributes to expression of naturally occurring mycoplasmosis.     

Pathogenicity of fowl adenovirus isolated from gizzard erosions to immuno-suppressed chickens

Pathogenicity of a fowl adenovirus (FAV), JM1/1 strain of serotype 1 derived from gizzard erosions of a broiler chicken, was examined to specific pathogen-free (SPF) chickens pre-treated with infectious bursal disease viruses (IBDVs) or cyclophosphamide (CY). Virulent IBDVs, classical type, were inoculated orally at 3 days of age of SPF chickens. CY was treated subcutaneously for 3 days after hatch. FAV was given orally at 30 days of age. At 40 days of age, all chickens were bled and autopsied for serology and gross observation. Gizzard lesions were ranked by the scores depending on their severities. IBDV- or CY-treated chickens showed significantly higher gizzard lesion scores than non treated birds. There were no gross lesions in any other organs except for bursal atrophy. Serologically, antibody production against FAV was highly suppressed by IBDV infection or CY treatment.     

Outbreak of herpesviral conjunctivitis and respiratory disease in gouldian finches

An outbreak of tracheitis, sinusitis, and conjunctivitis, originating in recently imported birds, caused high morbidity and mortality in a flock of finches in Central Illinois. Although several species were present, Gouldian finches (Erythrura [Chloebia] gouldiae) were most commonly and severely affected. Birds submitted for necropsy displayed microscopic lesions characteristic of herpesviral infection, including epithelial cytomegaly and karyomegaly with basophilic, intranuclear inclusion bodies in the nasopharynx, sinuses, trachea, parabronchi, conjunctiva, and occasionally the lacrimal gland or proximal proventricular glands. Viral particles consistent with herpesvirus were visualized within affected epithelial cells with electron microscopy. Based on a partial sequence of the viral DNA polymerase gene, this virus was found to be identical to a herpesvirus previously implicated in a similar outbreak in Canada and is most likely an alphaherpesvirus.     

Experimental rabies in skunks: mechanisms of infection of the salivary glands.

Striped skunks (Mephitis mephitis) were inoculated into the right submandibular salivary gland with street rabies virus. They were killed at various times after inoculation and several tissues were examined by immunofluorescence and light microscopy. Right and left superior cervical, nodose and trigeminal ganglia, medulla oblongata and at least three regions of right and left submandibular salivary glands were examined by the fluorescent antibody technique. Intracerebral titrations of salivary gland suspensions were made in weanling white Swiss mice. Immunofluorescent material (inoculum) was detected in septa and connective tissue surrounding secretory units of the right submandibular gland immediately after inoculation, but otherwise antigen was not detected in either right or left submandibular glands without coincident antigen in the medulla oblongata. This occurred first on day 12 in areas of the gland remote from the inoculation site. Titers of virus were low at this time. Serum neutralizing antibodies occurred by day 7 in a few skunks. The time of development and distribution of antigen strongly suggest that, even after direct inoculation, neural networks are necessary for development of widespread infection of the salivary gland. The early occurrence of serum neutralizing antibodies in some of the skunks suggests that the immune response was activated by virus in the inoculum since immunofluorescence was not detected in any tissue at this time.