Chick Embryo Passage Research Articles

Complete genomic sequence of a duck enteritis virus attenuated via serial passage in chick embryos.

Here, we present the complete genomic sequence of an attenuated duck enteritis virus (DEV). The Chinese standard challenge strain of DEV (DEV CSC) was serially passaged 20 times in chick embryo fibroblasts and then 85 times in chick embryos. The virus was attenuated and was avirulent to 2-month-old ducks. The attenuated DEV genome is 162,131 base pairs (bp) in length and as long as the parental genomic sequence. There are only 22 nucleotide substitutions, resulting in single amino acid changes in open reading frames LORF5, LORF4, UL41, UL39, UL32, UL13, UL10, UL3, US3, US4 and US7. The genome sequence has been deposited in the GenBank database under accession number KU216226. This study provides genetic information about DEV attenuation and further advances our understanding of the molecular basis of DEV pathogenesis.

Improved methods for isolation of avian influenza virus

Isolation of viruses using chick embryos is a classical virological method. Inoculation of the allantoic cavity and use of allantoic fluid is a common method of passaging isolated avian influenza viruses. In the present study, 2490 fresh fecal samples and 4967 old fecal samples were investigated and subjected to conventional passaging (allantoic fluid method). Two newly developed methods-the allantochorion and allantoic fluid mixed method and the chick embryo and allantoic fluid mixed method-were also examined. The rates of influenza virus isolation for these three methods were compared. There appeared to be little difference among these methods when fresh fecal samples were used. However, for the old fecal samples, isolation rates for influenza virus were significantly higher for the chick embryo and allantoic fluid mixed method compared with the conventional allantoic fluid method. All viruses isolated using the conventional allantoic fluid method were isolated successfully using the two newly developed methods. These results suggest that using chick embryos in conjunction with allantoic fluid is effective for early virus isolation, especially for fecal samples that are not fresh. Additionally, practical chick embryo passage methods are described that improve significantly the rate of isolation of influenza viruses from fecal samples of migratory birds in a complex wild ecological environment.

Plaque morphology of some South American strains of foot-and-mouth disease virus and the effects of polyionic compounds on plaque formation

The plaque forming ability of attenuated and field strains of FMDV of types A and O Vallee and C Waldmann were compared in BHK cell cultures. It was found that the vaccine strains attenuated by chick embryo passage produced smaller plaques. Reactor viruses recovered from vaccinated animals generally produced larger plaques than the vaccine and field strains. Type O vaccine, reactor and field viruses produced plaques of less variation in size and morphology. Field viruses of types A and C produced non-clear plaques which are readily distinguishable from the corresponding vaccine and reactor viruses.

The culture of dysentery bacillus in chick embryo

Shigella dysenteriae (type Shiga) has been cultured in chick embryo using Tanaka's method in which a hen's egg is settled in a longitudinal, standing position, that is, with its dull point up and its sharp one down.After several chick embryo passage uses the virulence of Shigella dysenteriae increased in the chick embryo, and then it increased in mice.It has been demonstrated that after several passage uses in certain species of animals, virulence increased in other species of animals as well as the passage animal.The above-mentioned fact explains the mechanism of some epidemic infections.

The Effect of Scald - Water Temperature on the Histologic Appearance of Chicken Skin

1. Adler, H. E. A comparison of some characteristics of Mycoplasma var. mycoides and Mycoplasma gallisepticum. Am. J. Vet. Res. 25, 243-245, 1964. 2. Hofstad, M. S., and L. Doerr. A chicken meat infusion medium enriched with avian serum for cultivation of an avian pleuropneumonia-like organism, Mycoplasma gallinarum. Cornell Vet.) 46 (3), 439-446. 3. Jungherr, E. L., R. E. Luginbuhl, and R. E. Jacobs. Pathology and serology of air sac infection. Proc. Book AVMA, 303-311, 1953. 4. United States Department of Agriculture, Agricultural Research Service. Standard methods for testing avian sera for the presence of Mycoplasma galliselpticum antibodies. Pamphlet. 5. van Herick, E. L., and M. D. Eaton, An unidentified pleuropneumonialike organism isolated during passage in chick embryos. J. Bact. 50, 47-55, 1945.

Antigenic Relationship of Toxoplasma gondii and Besnoitia jellisoni

Morphologically Besnoitia and Toxoplasma resemble each other. Immunologically they are different but appear to have certain antigens in common. Rabbits were infected with Toxoplasma gondii and Besnoitia jellisoni both of which had been passed through chick embryo. Dye tests and hemagglutination tests were done using both Toxoplasma and Besnoitia as antigens. The organisms for the dye test and for preparation of HA antigens were derived from peritoneal exudates of intraperitoneally infected mice. The dye test was specific for both organisms with no titer demonstrated in the heterologous system. On the other hand, cross-reactions were demonstrated between Toxoplasma and Besnoitia in the hemagglutination test. The homologous titer was always higher than the heterologous titer. Complement-fixation test using only the Toxoplasma antigen showed a cross-reaction with Besnoitia antisera from these rabbits. Agar-gel diffusion on micro slides revealed precipitin lines in common with both systems. Rabbits infected with Besnoitia showed no immunity when challenged with a virulent strain of Toxoplasma and succumbed as rapidly as controls not previously infected. On the other hand, rabbits surviving infection with an avirulent Toxoplasma strain were immune to challenge with virulent Toxoplasma. Frenkel (1960) reviewed a number of organisms resembling Toxoplasma morphologically. The trophozoites of Besnoitia jellisoni are so similar to Toxoplasma gondii that it would be difficult to distinguish the two organisms with certainty in a mixed population. Goldman et al. (1957), using a silver protein stain, found similar internal structures and reproductive stages in Besnoitia and Toxoplasma trophozoites. Besnoitia cysts are larger than those of Toxoplasma and also have a thicker wall lined by large nuclei (Frenkel, 1955). From previous work it appears that Besnoitia and Toxoplasma are immunologically distinct. Frenkel (1953) using the dye test, found no cross-reactions between the two organisms. Goldman et al. (1957) reported that fluorescein-labeled Toxoplasma antibody did not stain Besnoitia under conditions in which Toxoplasma stained brightly. Although no protozoan parasites have been conclusively incriminated in producing Toxoplasma dye test antibodies, no studies including hemagglutination (HA) antibodies have been reported. Received for publication 28 July 1964. MATERIALS AND METHODS New Zealand white rabbits weighing approximately 2 kg were inoculated intradermally (id) with 10,000 relatively avirulent Toxoplasma harvested from the chorioallantoic membrane of chick embryos infected via the yolk sac 7 days earlier. Rabbits Nos. 1069 and 1070 were inoculated with the Beverley strain and rabbit No. 1071 was inoculated with strain 113-CE. Both of these strains had been maintained by weekly passage in chick embryo for over a year. One rabbit (No. 1066) had been inoculated id with 5,000 Toxoplasma trophozoites of the virulent RH strain, and received Spiramycin 125 mg/kg subcutaneously for 2 weeks. Chemotherapy was necessary to prevent death which would otherwise have occurred in 7 to 10 days. Besnoitia jellisoni was received from Dr. J. K. Frenkel (University of Kansas Medical Center). Prior to passage through chick embryo for the present study it had been maintained in mice by intraperitoneal (ip) passage twice a week for 3 months. Two rabbits weighing 2 kg each were given an initial id inoculation of 10,000 Besnoitia trophozoites from chick embryo followed by a second inoculation of 10,000 organisms id 2 months later. All rabbits were bled for serological studies 2 to 3 months after infection. These rabbits plus two normal rabbits were then challenged id with 1,000 Toxoplasma of the virulent RH strain. They

CHANGES IN MOUSE PNEUMONITIS AGENT ASSOCIATED WITH DEVELOPMENT OF RESISTANCE TO CHLORTETRACYCLINE.

Moulder, James W. (University of Chicago, Chicago, Ill.), Dorothy L. Novosel, and Ilse I. E. Tribby. Changes in mouse pneumonitis agent associated with development of resistance to chlortetracycline. J. Bacteriol. 89:17-22. 1965.-A chlortetracycline-resistant mutant of mouse pneumonitis agent, a member of the psittacosis group of microorganisms, differed in several ways from the susceptible stock from which it had been derived by serial chick-embryo passage in the presence of the antibiotic. It was almost completely resistant to chlortetracycline, had a longer growth cycle, produced many more very large particles, was not neutralized by parent antiserum and vice versa, was 10 times as resistant to d-cycloserine, and was highly lethal to mice when inoculated intracerebrally. These changes probably do not occur independently of one another but are the reflection of some genetically controlled change in the surface structure of the mouse pneumonitis particle which enables the resistant mutant to multiply in the presence of chlortetracycline.

Nonpathogenic Rickettsias Related to the Spotted Fever Group Isolated from Ticks, Dermacentor Variabilis and Dermacentor Andersoni from Eastern Montana

Summary Forty strains of a rickettsia, referred to as the EM rickettsia, which was avirulent for guinea pigs and which retained this property after serial passage in chick embryos were isolated from Dermacentor variabilis collected in two counties of eastern Montana. The EM rickettsia possessed properties common to agents of the spotted fever group. However, although it appeared to be a variant of Rickettsia rickettsii, it could not be identified with any of the four previously distinguished type strains of this species. The EM rickettsia was also isolated from Dermacentor andersoni collected in one of these localities. EM isolates were compared with an avirulent U strain of R. rickettsii which had been recovered from D. andersoni from the Bitterroot Valley, Montana. They differed primarily in their respective capacities to produce cross-reacting antitoxin against the virulent R strain of R. rickettsii. Guinea pigs injected with 104 doses of EM rickettsias developed high titers of CF antibody but little, if any, antitoxin, against the R strain. In contrast, the U strain produced high titers of both antitoxin and CF antibody for the R strain. Guinea pig hyperimmune serum against the EM rickettsia neutralized toxins of three strains of R. rickettsii representing the R, S, and T types, respectively, but did not neutralize toxins of Rickettsia conori, Rickettsia siberica, or the rickettsia from the tick Amblyomma maculatum. A high rate of infection with EM strains was found in the tick populations examined. This rickettsia was recovered from about 6 to 8% of D. variabilis from each of five localities studied. With one exception all strains isolated in eastern Montana were of the EM type. One isolate which possessed the characteristics of strains carried by the tick Haemaphysalis leporis palustris was isolated from a lot of ten D. variabilis from the eastern Montana area. The T strain commonly isolated from D. variabilis from eastern United States was not encountered in the study area.

A host-induced modification of hemagglutinating virus of Japan (HVJ, Sendai virus) in its hemolytic and cytopathic activity

Host-induced changes of HVJ in hemolytic activity and in growth characteristics in mouse lung cells were investigated. HVJ grown in the allantoic cavity of the chick embryo is hemolytic, but the virus loses this property after a single passage in the mouse lung or after a single growth cycle in cultivated mouse lung cells. This nonhemolytic virus becomes hemolytic after a single chick embryo passage. Similar changes in virus growth characteristics were also observed. When HVJ was adapted to growth in monolayers of mouse lung cells, virus was produced within 24 hours of infection, but cytopathic effects were delayed, often for several days. After one passage in the chick embryo, however, this virus reproduced in the tissue culture with concomitant destruction of the cells. After a single passage in tissue culture the cytopathic virus returned to the noncytopathic condition.

Influenza Virus-Erythrocyte Interaction

Summary Early (3rd) and later (10th) chick embryo passage NA strain of influenza A virus adsorbs on sheep RBC at 4°C and elutes from these cells at 20°C at identical rates. Adsorption of virus is accompanied by agglutination of the RBC. When elution takes place at 20°C, disagglutination of RBC occurs earlier and proceeds faster with 3rd passage than with 10th passage NA virus. After inactivation of the virus enzyme by heat, the 10th passage virus fails to elute at 26°C, whereas the 3rd passage virus still dissociates promptly. Under these conditions RBC disagglutinate rapidly in samples containing 3rd passage virus but remain agglutinated in the presence of 10th passage virus. It is concluded that the binding forces which operate between early passage NA virus and sheep RBC are weak. On adaptation to a new host, i.e., the chick embryo, the virus undergoes a gradual change resulting in increased strength of these forces.

Mutations in pathogenicity of some recently isolated strains of fowl-plague virus

Summary Two out of four recently isolated field strains of fowl-plague virus failed to yield recognizable mutants during serial cultivation in pigeon and chick embryos. One strain became avirulent for fowls at the 11th pigeon-embryo passage, but capacity to infect fowls was completely or almost completely, lost during the next three passages in either chick or pigeon embryos, and was not regained until 27 to 30 additional passages had been made in chick embryos. The regained capacity of this strain to infect fowls was unaffected by further serial repass age in pigeon embryos. Other lines of this strain, started from material stored before any mutation had occurred, behaved differently under serial cultivation in pigeon embryos. One line remained lethal to rather more than 50 per cent of inoculated birds, while the virulence of a second line fell to a level at which it caused a mortality of about 10 per cent. Of two lines, derived from the series in which infectivity for fowls had been lost and reacquired, one showed an exaltation of virulence after further serial pigeon-embryo passage. A substrain “B” of this line, which had been passaged six times in chick embryos simultaneously with Newcastle-disease virus, was of remarkably low virulence and high antigenic capacity for fowls from that point onwards. The second line, subjected to 125 serial passages in the pigeon embryo was at that stage lethal to about 5 per cent of inoculated fowls. A subs train derived from one of the pigeon-embryo passaged strains by serial intracerebral passage in mice showed greatly reduced capacity to infect and immunize fowls. After 7 repassages in the pigeon embryo there was a sudden access of virulence for fowls. When the first passage in chick embryos of a fourth field strain. Of virus was stored at −20°C for 64 days, and then passaged in chick embryos, material from two successive passages immunized birds without causing any illness; but its full virulence was restored after a further 6 passages in chick or pigeon embryos. After a further 19 passages the virulence showed signs of declining. Virus stored in the original brain suspension for 88 days was still fully virulent after a single passage in chick-embryos, but showed signs of declining after 6 further passages in chick or pigeon-embryos. Mutual interfence was demonstrable in birds inoculated with mixtures of “attenuated” fowl-plague and Newcastle-disease viruses, but when titres of the two viruses were adjusted so that neither virus had a marked advantage in numbers most of the birds developed immunity to both diseases.

Poliomyelitis. II. Propagation of MEFI strain of poliomyelitis virus in developing chick embryo by yolk sac inoculation.

Summary1. The MEFl-A strain of human poliomyelitis virus(10), after 119 serial passages in suckling hamsters, has been maintained in one passage series for 41 consecutive generations and in a second series for 38 generations in the developing chick embryo by the yolk sac route of inoculation. Both lines of passages are still in progress. 2. The identity of the chick embryo adapted virus with human poliomyelitis virus was established by mouse neutralization tests in which anti-Lansing type immune sera from 3 different sources were used; by the ability of the passage virus to stimulate the formation of antibodies in monkeys which neutralize known virulent strains of Lansing type poliomyelitis virus; and by the ability of monkeys to withstand intracerebral challenge with known virulent strains of poliomyelitis virus after they had survived inoculation with the chick embryo passage virus. The identity of the passage virus was further established by its ability to induce typical symptoms and histopathologic le...

Antigenic similarity between the first and later egg passage strains of freshly recovered influenza A viruses.

SummaryCross-neutralization experiments in ovo with 1st and later chick embryo passage strains of freshly recovered influenza A viruses showed no definite difference in their antigenic composition. However, the reactivity relative to guinea pig and chicken erythrocytes underwent a rapid and profound change between the 1st and 2nd embryo passages.

The Isolation in Eggs of a New Filtrable Agent which may be the cause of Bovine Lumpy Skin Disease

SUMMARY: Using a filtrate from an emulsion of skin nodules, lymph node and a milk-duct nodule from a calf dead of bovine Lumpy Skin Disease as starting material, a filtrable agent was isolated by serial passage in chick embryos. The agent affects mainly the skin structures of the chick embryo and results in a characteristic shrunken featherless embryo tightly wrapped in its amnion, and with almost complete disappearance of amniotic fluid. The agent appears to be a filtrable virus approximately 10-25 mμ in diameter. It was not pathogenic for day-old chicks and adult fowls. The virus was neutralized by the serum of fowls which had received repeated intramuscular injections of the virus, suggesting that the virus is etiologically related to Lumpy Skin Disease in cattle. Further work is needed before such a relationship can be regarded as established. The virus withstood heating to 60° for 20 min., survived in 50% glycerol for at least 1 week, and at a temperature of +4° for at least a month. It was destroyed by boiling for 2 min. and by incubation at 37° overnight. Desiccation destroyed a large part of the virus in a suspension, but that part which withstood the drying process survived for at least a month.

THE PRODUCTION OF A PERSISTENT ALTERATION IN INFLUENZA VIRUS BY LANTHANUM OR ULTRAVIOLET IRRADIATION

The rates of elution from RBC of the Lee and PR8 strains of influenza virus were studied by means of a step-wise elution technique. By means of a single treatment with lanthanum acetate or irradiation with ultraviolet and subsequent passage in chick embryos, it was possible to alter the elution rate of the Lee strain so that it was similar to that of the PR8 strain. This alteration proved to be persistent on serial passage in the absence of the agent which caused it. As far as was determined, the elution rate of the virus appeared to be the only property which was altered. The phenomenon can be most readily understood on the assumption that the difference in elution rates of the two strains is due to a heterogeneous population of virus particles in the Lee strain with respect to elution rate.

ISOLATION OF MUMPS VIRUS FROM HUMAN BEINGS WITH INDUCED APPARENT OR INAPPARENT INFECTIONS

Exposure of fifteen children to mumps virus of fifth amniotic passage in chick embryos led to involvement of the salivary glands in six, orchitis in the absence of other manifestations of mumps in one, and to no signs of illness in eight. Attempts to isolate virus from the saliva of these individuals gave the following results: 1. All patients with involvement of the salivary glands excreted virus beginning on the 11th to 15th day after exposure, 2 to 6 days prior to onset of clinical signs of disease and extending up to the 4th day of illness. 2. The patient with primary orchitis without any recognized involvement of the salivary glands excreted virus for 2 days, beginning on the 15th day after exposure and 10 days prior to his illness. 3. Six of the eight children classified as having inapparent infections because of their serologic response in the absence of clinical signs of illness, began to excrete virus on the 15th to 16th day after exposure for from 1 to 9 or more days. The epidemiologic significance of these data is discussed.

Direct Isolation of Human Influenza Virus in Chick Embryos

Summary Of the filtrates from 54 throat washings tested by the chick embryo passage method 28 gave evidence of containing influenza virus. The strains were isolated directly in chick embryos, and the presence of hemagglutinins in the allantoic fluid was used as evidence for the presence of influenza virus. In most instances where a successful isolation was obtained, the positive result was first noted in the 2nd embryo passage. Inoculation of strains that had been isolated by this method, into mice, ferrets, and chick embryos, showed their capacity to infect ferrets and embryos in high titer, but they possessed a very low order of virulence for mice.

Characteristics of a Fixed Rabies Virus Cultivated on Developing Chick Embryos

SummaryAfter 47 passages through the developing chick embryo, rabies fixed virus shows no enhanced virulence for the chick embryo, as judged by the complete development of the embryo, the relatively low virus titre in the embryo brain and the scanty pathological changes in the brain. The virulence of the chick embryo passage virus for mice and guinea pigs remained unchanged, while that for rabbits appeared reduced considerably, as demonstrated by the prolonged incubation period, and duration of the infection. The specific anti-genic character of the chick embryo virus remained unchanged during 47 passages over a period of 18 months.