Various Routes Of Inoculation Research Articles

The impact of the experimental route of challenge on the host responses and pathogenesis of the Canadian Delmarva (DMV/1639) infectious bronchitis virus infection in laying chickens

Infectious bronchitis virus (IBV) infection can be associated with respiratory, renal, and/or reproductive diseases in chickens. Under natural conditions, conjunctiva, mucosa of upper respiratory tract, and cloaca are the main routes of IBV entry. Experimentally, the study of IBV infection involved various routes of inoculation. This study investigated the impact of adding the trachea as a potential route of viral entry to the oculo-nasal infection on the host responses, pathogenicity, and tissue tropism of the Canadian IBV Delmarva (DMV/1639) strain in laying chickens. Specific-pathogen-free laying chickens were divided into three experimental groups: control group (Con group), oculo-nasal challenged group (ON group), and oculo-nasal/intratracheal challenged group (ON/IT group); all groups were observed for 12 days post-infection (dpi). The clinical signs and reduction in egg production in the ON/IT group started slightly earlier compared to the ON group. At 12 dpi, the gross lesions in the ON/IT group were confined to the ovary, while the ON group showed regressed ovary and atrophied oviduct. Only the ON group showed significantly higher microscopic lesion scores in the lung, kidney, magnum, and uterus compared to the control group at 12 dpi. The oviduct tissues of the ON group showed a significant increase in B cells infiltration compared to ON/IT and control groups. The viral shedding (detected by quantitative reverse transcription polymerase chain reaction (qRT-PCR)), tissue tropism (detected either by qRT-PCR or immunohistochemistry (IHC)), T/natural killer cells infiltration in reproductive tract (detected by IHC), and antibody-mediated immune responses (measured by enzyme-linked immunosorbent assay) showed similar patterns in the ON and ON/IT groups.

Contemporary Circulating Enterovirus D68 Strains Infect and Undergo Retrograde Axonal Transport in Spinal Motor Neurons Independent of Sialic Acid.

Enterovirus D68 (EV-D68) is an emerging virus that has been identified as a cause of recent outbreaks of acute flaccid myelitis (AFM), a poliomyelitis-like spinal cord syndrome that can result in permanent paralysis and disability. In experimental mouse models, EV-D68 spreads to, infects, and kills spinal motor neurons following infection by various routes of inoculation. The topography of virus-induced motor neuron loss correlates with the pattern of paralysis. The mechanism(s) by which EV-D68 spreads to target motor neurons remains unclear. We sought to determine the capacity of EV-D68 to spread by the neuronal route and to determine the role of known EV-D68 receptors, sialic acid and intracellular adhesion molecule 5 (ICAM-5), in neuronal infection. To do this, we utilized a microfluidic chamber culture system in which human induced pluripotent stem cell (iPSC) motor neuron cell bodies and axons can be compartmentalized for independent experimental manipulation. We found that EV-D68 can infect motor neurons via their distal axons and spread by retrograde axonal transport to the neuronal cell bodies. Virus was not released from the axons via anterograde axonal transport after infection of the cell bodies. Prototypic strains of EV-D68 depended on sialic acid for axonal infection and transport, while contemporary circulating strains isolated during the 2014 EV-D68 outbreak did not. The pattern of infection did not correspond with the ICAM-5 distribution and expression in either human tissue, the mouse model, or the iPSC motor neurons.IMPORTANCE Enterovirus D68 (EV-D68) infections are on the rise worldwide. Since 2014, the United States has experienced biennial spikes in EV-D68-associated acute flaccid myelitis (AFM) that have left hundreds of children paralyzed. Much remains to be learned about the pathogenesis of EV-D68 in the central nervous system (CNS). Herein we investigated the mechanisms of EV-D68 CNS invasion through neuronal pathways. A better understanding of EV-D68 infection in experimental models may allow for better prevention and treatment strategies of EV-D68 CNS disease.

Recovery of Campylobacter from External and Internal Spleen Samples from Baby Broiler Chicks Following Various Routes of Inoculation

AbstractCampylobacter have been found in lymphoid tissue of poultry, but the location in these tissues has not been determined. The objective of this study was to determine if Campylobacter are on the outside or on the inside of the spleen. For external (ES) spleen sampling 2 days post‐inoculation, each spleen was rinsed with 3 mL of Bolton's enrichment broth (BEB). For internal (IS) spleen sampling, each rinsed spleen was submerged into 70% ethanol for 10 s, removed and submerged into 0.85% saline. The spleen was then placed into a sterile bag, macerated with 3 mL of BEB and stomached for 30 s before incubation. All samples and the correlating ceca were sampled for Campylobacter jejuni. Overall in four different experiments with baby chicks inoculated by oral, ocular or cloacal routes, Campylobacter was recovered from 75% (82/109) of the ES, 71% (77/109) of the IS and from 100% (109/109) of the ceca. Ethanol and saline samples were all negative for Campylobacter, suggesting that ethanol immersion is adequate to kill C. jejuni on the external surface. With Campylobacter in the internal tissue of the spleen, the organism appears to be systemic. Further research will determine the colonization mechanisms required for this bacterium in these different tissues.Practical ApplicationsIt is very difficult to sample the free flowing blood of a living chicken for bacteria without introducing contamination from the skin. This paper describes an effective method of sampling the internal portion of the spleen without introducing external contamination. By doing so, this indicates that the bacteria were systemic and will travel with the blood to all of the internal organs and tissues throughout the bird's body. Therefore when the bacteria are not isolated from certain tissues, it means that reservoirs were either not established or did not persist in that particular tissue even though the bacteria did flow through with the blood. Unless all of the potential reservoirs and sources of bacteria such as Campylobacter and Salmonella are identified and fully understood, successful intervention strategies may not be possible. It would be important to know if reservoirs of these bacteria present in internal organs and tissues can periodically seed the intestinal and/or reproductive tracts of adult breeder birds, thus providing a source of contamination to the offspring.

Zosteriform spread of Herpes simplex virus 1 as a model to study the role of leukocyte subpopulations in the mortality induced during acute infection (P4373)

Abstract Immune response against Herpes simplex virus 1 (HSV-1) has been studied by various routes of inoculation in models of herpes. Zoteriform spread of HSV-1 infection occurs after primary inoculation of the skin of mice. This inoculation route has been used for study the pathogenesis of infection at nervous system level but little is known about the mechanism of immune response in this model. The aim of this work is to study the infiltrated of leukocytes in the inguinal lymph nodes (ILN) and correlate changes in leukocyte subpopulations with mortality in BALB/c mice cutaneously infected with HSV-1. We observed that infected mice showed 70-100% mortality during the acute infection. We removed inguinal and mesenteric lymph nodes (MLN) from infected mice and analysed phenotype of CD3, CD4, CD8, and CD19 cells from 0 to 15 days post-infection. The percentage of CD19+ cells and CD3+ cells in the ILN from infected mice changed from 20% to 50% and from 70% to 40% respectively. In contrast, such changes were not observed in MLN, suggesting that during acute infection immune response is elicited only at the region of the infection. Kinetic analysis of CD4+ and CD8+ T cells showed that the HSV-1-infection induced the expression of CD11c in T cells. Our findings show that HSV-1 infected mice retain a CD11c+ leukocyte infiltrate that could be involved in mortality. The role of CD11c+ cells will be further studied using immune-modulators that modify the percentage of those populations.

Effect of Different Routes of Inoculation on Infectivity and Viral Shedding of LPAI Viruses in Mallards

We studied the effect of different routes of inoculation on the infectivity and duration of viral shedding in mallards (Anas platyrhynchos) infected with wild bird-origin low pathogenic avian influenza viruses (LPAIVs). One-month-old mallards were inoculated with 10(6) median embryo infectious doses of either A/mallard/MN/199106/99 (H3N8) or A/mallard/MN/355779/00 (H5N2) via 1 of 5 different routes: intranasal (IN), intratracheal (IT), intraocular (IO), intracloacal (IC), or intra-ingluvial (II). Birds in all routes of inoculation groups became infected with LPAIV as detected by virus isolation, real time reverse transcription polymerase chain reaction, and serology. Mallards in different route of inoculation groups had similar viral shedding through oropharynx and cloaca from 1 day postinoculation (dpi). The peak of oropharyngeal (OP) viral shedding was reached between 2 and 3 dpi in all routes of inoculation groups infected with either virus. The peak of cloacal (CL) viral excretion was reached between 2 and 3 dpi in all routes of inoculation groups infected with H3N8 LPAIV and in the IO-, IC-, and II-inoculated groups infected with H5N2 LPAIV, with a delayed and shorter peak for the IN- and IT-inoculated groups. The birds inoculated via the II route had more productive OP and CL viral shedding after infection with either LPAIV, as evidenced by higher number of swabs testing positive over the study period. In conclusion, mallards can be infected with LPAIV by various routes of inoculation, and this corroborates their high susceptibility to infection by these viruses.

The Effect of Route of Inoculation on the Virulence of Raptorial Pasteurella multocida Isolates in Pekin Ducks (Anas platyrhynchos)

The purpose of this study was to determine the virulence of raptorial Pasteurella multocida for ducks and the effect of various routes of inoculation on virulence. Four-week-old Pekin ducks (Anas platyrhynchos) were challenged with one of three raptorial isolates (RTHA-2, RTHA-4, or WESO-1) by one of five inoculation routes (intranasal, intraocular, intravenous, oral, and subcutaneous). Ducks were monitored daily for mortality until 2 wk postchallenge. Results indicated that the intravenous route caused the most mortality for all isolates and that significant variation existed in the virulence among the sources of P. multocida, with WESO-1 causing the least mortality of the isolates tested.

Histological studies of bovine herpesvirus type 4 infection in non-ruminant species

The pathology of bovine herpesvirus type 4 (BHV-4) infection was studied in cats, rabbits and guinea pigs. Twenty kittens, twenty-two rabbits and ten guinea pigs, some treated with glucocorticoid were inoculated with a BHV-4 strain of feline origin, via various routes of inoculation (conjunctival, intranasal, peritoneal). Clinical signs were recorded. After euthanizing at different post inoculation days macro- and microscopic changes were observed by necropsy and in hematoxylin-eosin stained histological sections. The presence of the virus in organs was detected by immunohistochemistry and a nested PCR assay. Inclusion bodies and monoclonal antibody-stained cells were found in the conjunctiva, trachea, lungs, spleen and lymph nodes. Most of the lesions were localized to the respiratory and the immune system. The macro- and microscopic lesions and clinical signs were more severe in kittens and guinea pigs. The histological data indicated that cats, especially kittens, were susceptible for BHV-4 and the infection was not confined to the urinary bladder.

Use of life and inactivated Salmonella enteriditid=s phage type 4 vaccines to immunise laying hens against experimental infection

Four groups of Dekalb Delta commercial layer hens (8 hens per group) were reared individually in cages in an isolation unit. At the age of 8 weeks, groups 1 and 2 were vaccinated with 9R Salmonella gallinarum live (9R live) vaccine. At the age of 18 weeks, group 1 was re-vaccinated with 9R live S. enteritidis vaccine, while groups 2 and 3 were vaccinated with S. enteritidis bacterin. At the age of 22 weeks, groups 2 and 3 were re-vaccinated with S. enteritidis bacterin. Group 4 was not vaccinated and was kept as a control group. At 24, 27 and 30 weeks of age, the four groups were challenged using various concentrations of S. enteritidis phage type 4 and various routes of inoculation. Eggs, cloacal swabs and blood were collected weekly for bacteriological and serological examination. Isolation of the challenge organism from the eggshells and egg contents of the vaccinated groups was significantly lower than with the control group. There was no correlation between humoral antibodies and the shedding of the challenge organism in eggs or cloacal swabs. A combined vaccination programme of 9R live S. enteritidis vaccine and S. enteritidis bacterin provided better protection to laying hens than either vaccine administered alone. Vaccination should never be performed alone but in conjunction with other measures related to veterinary hygiene and good management.

Relative Infectivity of Borrelia Burgdorferi in Lewis Rats by Various Routes of Inoculation

Various routes of Borrelia burgdorferi infection were studied in laboratory rats. Three-week-old Lewis rats were inoculated either intradermally (i.d.), intraperitoneally (i.p.), or oronasally (o.n.) with serial 10-fold dilutions of B. burgdorferi. Thirty days later, groups of rats were killed and serology, splenic culture, and histology were used to evaluate infection. Rats were successfully infected i.d. with 10(2-4) organisms or i.p. with 10(4-5) organisms. Neither three-day-old nor three-week-old rats were successfully infected o.n. with up to 10(6) organisms. For contact transmission, three-day-old or three-week-old inoculated rats were housed with unexposed littermates for 30 days. Inoculated rats became infected but contact rats remained free of infection. To study in utero transmission, five pregnant female Lewis rats were inoculated i.p. with 10(6) spirochetes at four days gestation. Although adult females seroconverted or had positive splenic cultures at 20 days gestation, the placentas and fetuses were uniformly culture-negative. Venereal transmission from seven infected females or six infected males to uninfected rats of the opposite sex was not demonstrated.

センダイウイルス不活化ワクチン投与ラットにおける血清抗体の産生と母仔移行

Adult male and female rats were inoculated with tween 80-ethylether-formalin-ultraviolet inactivated Sendai virus (Sv-V) and examined for the production of hemagglutination inhibition (HI) antibody. There were no significant differences in the antibody titers between males and females, and among the various routes of inoculation except for the intranasal which was not effective. The antibody became detectable 7 days after a single inoculation with 10(5) HAU of Sv-V. The antibody titer, which had its peak 21 days after the inoculation, persisted for 200 days and declined gradually thereafter. The HI antibody titers were correlated with inoculated Sv-V doses and a predominant booster reaction with the vaccine was observed. Maternal antibodies were detected in sucklings born to dams hyperimmunized with the vaccine. The titers were similar to those of the dams until 3 weeks after birth but declined rapidly after weaning at 4-week-old. The titers of fetuses and neonates before suckling were significantly lower than those of the sucklings.

Pathogenesis of herpes simplex virus types 1 and 2 in mice after various routes of inoculation

The pathogenesis of herpes simplex virus (HSV) types 1 and 2 was compared after inoculation of mice by different routes. Intravaginal inoculation of HSV-1 and HSV-2 produced a local infection, with virus recovery from the vagina through 5 days. Virus was recovered from the spinal cords 4 to 5 days after inoculation but not from liver, kidney, lung, spleen, or blood. Intravenous or intraperitoneal inoculation of HSV-2 produced a focal necrotic hepatitis similar to that described previously (S. C. Mogenson, B. Teisner, and H.K. Andersen, 1974). The viral etiology of the liver lesions was confirmed by virus isolation (through 4 days) and electron microscopy. No evidence of infection of the kidney, lung, blood, or spleen was observed, although virus was isolated from spinal cord homogenates 7 days after inoculation. HSV-1 inoculation by the intraperitoneal or intravenous route resulted in virus isolation from the kidney during the 7-day harvest period, without producing overt pathological changes. Virus was isolated from spinal cord homogenates 2 to 3 days after HSV-1 inoculation but not from homogenates prepared from spleen, lung, or blood. Increases in serum transaminase activity were observed after systemic (intravenous) inoculation of HSV-2 but not after HSV-1 inoculation.

Studies on the malayan forest rat filaria, Breinlia booliati (Filarioidea: Onchocercidae): Transmission to laboratory rats

Breinlia booliati, a new filarial parasite of Rattus sabanus, has been successfully transmitted to 3 strains of laboratory rats by various routes of inoculation. Out of a total of 222 rats, 54 showed microfilaraemia with a mean prepatent period of 12·5 weeks. Out of 168 amicrofilaraemic rats necropsied at 20 weeks after inoculation, 67 harboured adult worms localized in the thoracic and abdominal cavities. Unisexual infections occurred frequently ( 58 67 rats). In bisexual amicrofilaraemic infections, adult male and female worms were recovered either in the same location or separately in the thoracic and abdominal cavities. Female rats were more susceptible to infection than male rats. The Charles River inbred rats and outbred albino rats were more susceptible than the HS (Hooded) inbred rats. This rat-filaria combination may provide a useful experimental model for studying various aspects of filariasis.

Teratogenic effects of Colorado tick fever virus in mice.

The teratogenic effects in mice of Colorado tick fever (CTF) virus, a human pathogen, were demonstrated. Various routes of inoculation, gestational periods, and dosages of virus were studied. The incidence of stillbirth and neonatal death increased significantly after inoculation of pregnant mice with CTF virus. CTF virus crossed the placenta and replicated to high titers in the placental and fetal tissues. Mice that survived the first six weeks of life had neutralizing antibody to CTF virus. Neutralization tests, in which pregnant mice were inoculated with both CTF virus and specific antiserum to CTF virus, validated the teratogenic effects of CTF virus in mice.

Experimental infections of lambs with Rickettsia prowazeki.

Weanling lambs were inoculated by intraperitoneal, intravenous, and subcutaneous routes with 105.3 or 107.3 egg 50% infectious doses of the ZRS strain of Rickettsia prowazeki. Control lambs were inoculated with identical amounts of the same infectious material inactivated with formalin. Tissue and blood samples from animals inoculated with infectious material were tested for the presence of rickettsiae by inoculation into embryonated eggs, guinea pigs, and meadow voles (Microtus pennsylvanicus) and by fluorescence microscopy with specific antiserum. Generalized rickettsial infections in lambs were not produced despite the use of various routes of inoculation. R. prowazeki was isolated in some instances from brain, lung, spleen, liver, and subcutaneous tissue 6 or 7 days after inoculation, but not 29 days after inoculation. Rickettsemia was not demonstrated under any of the conditions tested. Serological and clinical responses in lambs receiving infectious inocula were not distinguishable from corresponding responses in lambs that had received comparable amounts of formalin-killed inoculum. The pattern of isolations of R. prowazeki from various lamb tissues was consistent with a picture of local self-limited infections unaccompanied by rickettsemia. These findings are compared with those obtained from infected guinea pigs and are discussed with regard to the hypothesis of extrahuman reservoirs of R. prowazeki.

Further studies on the mode of transmission of herpes-like virus in guinea pigs.

SummaryHartley guinea pigs can be infected with HLV by various routes of inoculation. These animals were found to be uniformly susceptible to intranasal inoculation of 3.0 log TCID50 of virus. Infection could also be achieved by oral inoculation. Four out of 7 animals inoculated orally had virus in their blood. Contact infection was achieved in 2 out of 7 animals that were housed with or adjacent to infected ones. Incidence of virus infection in newborns from both intraperitoneally inoculated or naturally infected parents was low, but the frequency increased to 59% or higher when the offspring were 6 months of age or older. No virus was isolated from the offspring of intracerebrally inoculated guinea pigs.

Experimental infection of calves with a bovine adenovirus.

SummaryCalves were experimentally infected with a bovine adenovirus using various routes of inoculation. Respiratory illness was observed in calves inoculated either intranasally or intratracheally. The virus had no effect on calves infected via the eye. Control calves remained normal throughout the experiment. The virus was recovered principally from rectal swabs of infected calves for 7 days. All infected calves responded serologically to the virus.

COCCIDIOIDOMYCOSIS: STUDIES OF CANINE VACCINATION AND THERAPY.

Abstract : A three-phase study of vaccination and antibiotic therapy in experimental pulmonary coccidioidomycosis of dogs was made to determine: (a) the efficacy of various routes of inoculation of a formalin-killed, arthrospore vaccine; (b) the combined effects of vaccination and oral Amphotericin B therapy administered immediately following respiratory exposure to Coccidioides immitis; and (c) renal damage or nephrotoxicity resulting from oral Amphotericin B therapy. Neither of the pulmonary routes of vaccination (aerosol of intratracheal) provided protection against a subsequent respiratory challenge of approximately 80,000 C. immitis arthrospores, either singly or in combination with oral Amphotericin B therapy (150 milligrams per day for 20 days following challenge), nor did subcutaneous vaccination or therapy alone. However, eight of twelve dogs receiving both subcutaneous vaccination and therapy completely resisted the respiratory challenge; the remaining four exhibited very minimal, self-contained disease. Histopathological examination revealed no renal damage or nephrotoxicity in any of the dogs receiving Amphotericin B therapy (total dose in excess of three grams); their blood urea nitrogen levels remained within normal limits.

Cultivation of Infectious Laryngotracheitis Virus in Embryonated Chicken Eggs by Various Routes of Inoculation

ULTIVATION of infectious laryngotracheitis (ILT) virus in embryonated chicken eggs by Hitchner and White5 indicated that highest titers resulted from inoculation of the chorioallantoic membrane (CAM), prepared by the formation of an artificial air cell'.1 Chomiak et al.2 found that the chorioallantoic sac (CAS) method was as sensitive as the CAM method. Fabricant4 described a modified technique, designated the CCA method, in which the inoculum was deposited on the inner shell membrane through the air sac. The material was carried to the CAM by a single needle puncture. A small amount of the inoculum would be carried into the allantoic sac by this procedure. For standardization and improvement of techniques for the isolation of ILT virus from field cases, it was considered necessary to evaluate the various routes of inoculation. The presence of ILT virus is measured by the production of plaques on the CAM. Four methods of inoculation were employed, and the titers used to determine which method was the most effective. When plaque formation was not observed with the original inoculum, materials were collected for passage. Amnioallantoic fluid (AAF) and CA membranes were harvested from embryos inoculated by the various methods at intervals fol-

Antigenically modified red cells in chickens infected with Newcastle disease.

Summary1. Antisera specific for either NDV or influenza virus-treated chicken erythrocytes were prepared. 2. Virus modified red cells were detected in chickiens infected with NDV by various routes of inoculation by use of the specific antiserum. 3. Hemagglutinins for NDV-treated chicken red cells were also detected in the serum of infected chickens.

A pneumotropic virus isolated from C3H mice carrying the Bittner Milk Agent.

1. Five isolations of a virus which causes a fatal pneumonitis of suckling mice by various routes of inoculation have been made from C3H mice known to carry the Bittner Milk Agent. 2. Older suckling mice are increasingly resistant to this virus and no apparent illness has been produced by inoculation of adult mice. 3. Specific neutralizing antisera have been produced in rabbits. 4. Due to its unique behavior and failure to show serologic or other relationships to known mouse viruses, K-virus has been regarded as an hitherto undescribed agent. 5. Long term experiments have been set up to determine, if possible, whether K-virus is identical with or related to the Bittner Milk Agent.