An Adenovirus Isolated from the Feces of Mice

Experimental infection of an inbred strain of DK1 mice was carried out with a mouse adenovirus strain K87, isolated from the feces of apparently healthy DK1 mice. Strain K87 was orally administered to four-week-old mice and the virus was recovered from their feces for at least 3 weeks. The highest virus titers in the feces were observed between 1 and 2 weeks after inoculation. In 7-week-old mice the period of virus excretion was about one week shorter. When neonatal or 2-week-old mice were administered virus orally, somewhat irregular results but similar to those from the 4 or 7-week-old mice were obtained. In these infected mice, virus growth was detected in the intestinal tract but not in oropharyngeal washings, nasal tissue, lung, spleen or urine. After inoculation through several parenteral routes, the virus was also detected in the feces and virus growth seemed to be limited mainly to the intestinal tract. No clinical manifestations were observed in any infected mice. When the virus was almost undetectable in the mice infected either orally or parenterally, the virus was readministered orally, but no further virus was recovered in the feces. Neutralizing antibody in the serum was detected 3 weeks after primary inoculation. Induction of immunological tolerance was examined by inoculating mice in utero 2–4 days before birth, but no evidence of induced tolerance was obtained. The use of the adenovirus strain K87-mouse system as a model system for the study of the infectious process and immune mechanisms is suggested because strain K87 has a tissue tropism analogous to many human adenoviruses.     

Modification of low virulent Newcastle disease virus infection in chickens infected with reticuloendotheliosis virus

One-day-old SPF chicks were inoculated with reticuloendotheliosis virus (REV) which had been isolated from contaminated Marek's disease vaccine. Then they were subjected to super infection with the B1 or TCND strain of Newcastle disease virus (NDV) and examined for virus recovery, antibody response and the appearance of symptoms. Regardless of the time, from 0 to 8 weeks, of inoculation with the NDV-B1 strain after the REV infection, the antibody response was suppressed and the duration of the NDV recovery prolonged. Specific death preceded by severe respiratory or neural signs occurred more frequently to chicks inoculated with REV than to uninoculated controls after inoculation with the NDV-B1 strain in the neonatal stage or with the NDV-TCND strain at 5 weeks of age.     

Royal pastel mink respond variously to inoculation with Aleutian disease virus of low virulence.

Information was sought on the varied responses of royal pastel mink (a non-Aleutian genotype) to Aleutian disease virus of low virulence. Thus, of 20 yearling female pastel mink inoculated subcutaneously with a large amount of the Pullman strain of Aleutian disease virus, only 3 succumbed to the disease. Of the other 17 mink, 3 had neither viremia nor a rise in level of serum gamma globulin during the 24 weeks after inoculation. The other 14 mink were viremic for variable periods during the first 12 weeks. In only five mink was the viremia accompanied by elevated levels of serum gamma globulin, usually from week 8 on. Of the 16 subclinically infected mink that did not succumb to intercurrent disease and otherwise remained healthy, 9 were examined at 19 to 31 months for persisting virus. In only one mink, small amounts were detected in the mesenteric lymph node and spleen nearly 28 months after inoculation. The other seven mink that survived the infection were not protected when challenged 31 months later with a small amount of the highly virulent Utah-1 strain. Even though still poorly understood, these varied responses of the royal pastel mink to infection with Aleutian disease virus of low virulence have important pathogenetic and epidemiological implications.     

Tenotomy of the avian anterior latissimus dorsi muscle. II. Can regeneration from the stump occur in the pigeon?

Because the chick's anterior latissimus dorsi muscle (ALD) regenerates a fast-twitch muscular connection after tenotomy, the pigeon's ALD was tenotomized, either at the origin or through the muscle 0.5 cm from the origin, to determine whether this muscle behaves similarly to the chick muscle. These procedures were compared in pigeons operated upon at 7 weeks, versus 5 to 9 months of age. The pigeon's ALD did not regenerate a new connection, and other differences were observed between the pigeon and chick ALD. The pigeon ALD has only a single slow muscle-fiber type, has fewer fast fibers, and transforms to a fast-twitch muscle more readily than the chick ALD after tenotomy. The transformation of muscle fiber types occurred more readily in the older pigeons than those tenotomized at 7 weeks of age. Tenotomy induced morphological alterations of the muscle fiber structure in all of the pigeons, which is in contrast to the absence of changes in the tenotomized chick ALD. Therefore the pigeon and chick ALD respond completely differently to tenotomy.     

Electrophysiological properties of biventer cervicis muscle fibers of normal and roller pigeons.

Cable parameters, excitability characteristics, and contractile response to acetylcholine were measured in biventer cervicis muscles from Helmet pigeons, Racing Homer pigeons and Parlor (nonflying) Roller pigeons. Cable parameters for the three strains, were respectively: calculated diameter, 30.1, 42.5, and 37.3 mum; membrane resistance, 450, 556, and 386 omega-cm2; membrane capacitance, 4.2, 3.9, and 4.5 muF/cm2, and myoplasmic resistivity, 79, 185, and 116 omega-cm. Significant differences between excitability characteristics of Homer pigeon and Roller pigeon fibers were a 17% shorter maximal latency for spike initiation (P less than 0.025) and 24% lower rheobasic current (P less than 0.05) in Roller fibers. Dose-response curves of isolated biventer cervicis to acetylcholine revealed slight, but significant, differences between Helmets and Rollers. These are the first electrophysiological data from pigeon skeletal muscle and the first from any avian biventer cervicis. The biventer muscles of chickens contain mainly "slow" fibers, but our results show that pigeon biventer fibers have properties similar to the "fast" PLD fibers of the chicken. Furthermore, the existence of different myoplasmic resistivities for each strain of pigeons used in this study suggests the need for more careful determination of this parameter in electrophysiological investigations. Although our results show that Roller pigeon fibers differ from those of nonrolling pigeons in the respects described above, these differences are minor in comparison to the severe behavioral abnormalities of Roller pigeons. Some yet untested component of neuromuscular transmission may be directly involved in the rolling phenomenon, but the differences we report may simply be due to strain differences, muscle hypertrophy, or a more severe defect elsewhere in the nervous system.     

Serological Diagnosis of Classical Swine Fever

Antibody levels in post-infection sera from a pig inoculated with a low virulent strain of classical swine fever virus (Hannover 62) and in sera from two pigs inoculated with another low virulent strain (Spielbach 66) and from an in-contact pig were assayed by complement fixation and immunofluorescence using classical swine fever virus (ALD strain) and bovine virus diarrhoea virus (UG 59 strain) as antigens. The complement fixation test used was modified by addition of a preparation of porcine Glq to the complement and by mercaptoethanol treatment of the immune serum before use. The mercaptoethanol treatment of the immune serum resulted in complete elimination of a haemolytic prozone often seen with porcine immune sera. In the sera from the inoculated animals complement-fixing antibodies appeared earlier than neutralizing antibodies. A few weeks after inoculation there was a correlation between the presence of complement-fixing and neutralizing antibodies. During the entire observation period of 13 weeks it was not possible to demonstrate complement-fixing or neutralizing antibodies in serum from a pig exposed to infection by contact with the two pigs inoculated with the Spièlbach 66 strain of classical swine fever virus.     

Bluetongue virus: (1) in pregnant white-tailed deer (2) a plaque reduction neutralization test

Six white-tailed deer (Odocoileus virginianus) were infected with bluetongue virus (BTs, vaccinal strain) approximately one-third of the way through their gestation period. One deer died of bluetongue 21 days after inoculation. Of the five surviving the infection, one had two mummified fetuses, and the others no fetuses upon euthanasia two weeks after term. Fetuses were present in two control deer and in the one which died of bluetongue. A plaque reduction neutralization test for bluetongue virus was developed and described for the first time and its sensitivity illustrated by high post inoculation titers which ranged from 1:3200 to greater than 1:16000.     

Classical Swine Fever Virus vs. Classical Swine Fever Virus: The Superinfection Exclusion Phenomenon in Experimentally Infected Wild Boar

Two groups with three wild boars each were used: Group A (animals 1 to 3) served as the control, and Group B (animals 4 to 6) was postnatally persistently infected with the Cat01 strain of CSFV (primary virus). The animals, six weeks old and clinically healthy, were inoculated with the virulent strain Margarita (secondary virus). For exclusive detection of the Margarita strain, a specific qRT-PCR assay was designed, which proved not to have cross-reactivity with the Cat01 strain. The wild boars persistently infected with CSFV were protected from superinfection by the virulent CSFV Margarita strain, as evidenced by the absence of clinical signs and the absence of Margarita RNA detection in serum, swabs and tissue samples. Additionally, in PBMCs, a well-known target for CSFV viral replication, only the primary infecting virus RNA (Cat01 strain) could be detected, even after the isolation in ST cells, demonstrating SIE at the tissue level in vivo. Furthermore, the data analysis of the Margarita qRT-PCR, by means of calculated ΔCt values, supported that PBMCs from persistently infected animals were substantially protected from superinfection after in vitro inoculation with the Margarita virus strain, while this virus was able to infect naive PBMCs efficiently. In parallel, IFN-α values were undetectable in the sera from animals in Group B after inoculation with the CSFV Margarita strain. Furthermore, these animals were unable to elicit adaptive humoral (no E2-specific or neutralising antibodies) or cellular immune responses (in terms of IFN-γ-producing cells) after inoculation with the second virus. Finally, a sequence analysis could not detect CSFV Margarita RNA in the samples tested from Group B. Our results suggested that the SIE phenomenon might be involved in the evolution and phylogeny of the virus, as well as in CSFV control by vaccination. To the best of our knowledge, this study was one of the first showing efficient suppression of superinfection in animals, especially in the absence of IFN-α, which might be associated with the lack of innate immune mechanisms.     

Electrophysiological properties of biventer cervicis muscle fibers of normal and roller pigeons

Cable parameters, excitability characteristics, and contractile response to acetylcholine were measured in biventer cervicis muscles from Helmet pigeons, Racing Homer pigeons and Parlor (nonflying) Roller pigeons. Cable parameters for the three strains, were respectively: calculated diameter, 30.1, 42.5, and 37.3 m̈m; membrane resistance, 450, 556, and 386 ω · cm²; membrane capacitance, 4.2, 3.9, and 4.5 m̈F/cm², and myoplasmic resistivity, 79, 185, and 116 ω · cm. Significant differences between excitability characteristics of Homer pigeon and Roller pigeon fibers were a 17% shorter maximal latency for spike initiation (P < 0.025) and 24% lower rheobasic current (P < 0.05) in Roller fibers. Doseresponse curves of isolated biventer cervicis to acetylcholine revealed slight, but significant, differences between Helmets and Rollers. These are the first electrophysiological data from pigeon skeletal muscle and the first from any avian biventer cervicis. The biventer muscles of chickens contain mainly “slow” fibers, but our results show that pigeon biventer fibers have properties similar to the “fast” PLD fibers of the chicken. Furthermore, the existence of different myoplasmic resistivities for each strain of pigeons used in this study suggests the need for more careful determination of this parameter in electrophysiological investigations. Although our results show that Roller pigeon fibers differ from those of nonrolling pigeons in the respects described above, these differences are minor in comparison to the severe behavioral abnormalities of Roller pigeons. Some yet untested component of neuromuscular transmission may be directly involved in the rolling phenomenon, but the differences we report may simply be due to strain differences, muscle hypertrophy, or a more severe defect elsewhere in the nervous system.     

Diagnosis of bovine respiratory syncytial virus infection by complement fixation test.

The complement fixation test by the microtiter method was applied to the serological diagnosis of bovine respiratory syncytial (RS) virus infection. When used as complement fixing antigens, untreated infected cell culture fluid, fluorocarbon-treated, and ether-treated materials showed no differences in antigenicity among them. The complement fixing antigenicity of bovine RS virus appeared in bovine kidney and Vero cell cultures for the first time 4 days after inoculation. Both the infectivity and complement fixing antigenicity reached a maximum 6 days after inoculation. In detecting complement fixing antibody from infected cattle, the most outstanding specific reaction was obtained when 5% fresh normal calf serum had been added to the diluent of complement. Neutralizing and complement fixing antibodies were examined in serum samples from two cattle in the course of experimental infection. It was found that both antibodies turned to be positive 2 weeks after inoculation. There was a linear correlation between neutralizing and complement fixing antibody titers, when serum samples from 40 natural cases were tested in the acute and convalescent stages. In addition, common antigenicity was demonstrated between the virus of bovine origin and the Long strain of human RS virus by complement fixation test.     

First report of establishment of Trypanosoma evansi infection in pigeon nestlings (Columba livia)

Trypanosomosis (surra) caused by Trypanosoma evansi is quite common among horses where the parasite is endemic. In the present study, T. evansi was isolated from an infected horse and maintained by subinoculation in Swiss albino mice. At the peak of parasitemia (5 x 10(6) parasites per ml of blood), 0.25 ml of the tail blood from infected mice was inoculated intraperitoneally and subcutaneously to 2 groups of adult pigeons and 2 groups of pigeon nestlings. Four days after inoculation, the trypanosomes occurred in the peripheral circulation of pigeon nestlings, but no parasitemia was observed in adult pigeons. The body temperatures of infected nestlings increased to 104 F, whereas uninfected controls remained steady at 102 F; thus, elevated temperatures coincided with parasite presence in the peripheral circulation. A decrease in hemoglobin concentration of blood also was observed in infected nestlings. On microscopic examination, increases in length and breadth of trypomastigotes and vigorous flagellar movement of the parasites were observed. The virulence and pathogenicity of the parasites after adaptation to nestlings remained unchanged for albino mice as proved by the death of all subinoculated mice. Furthermore, polymerase chain reaction studies confirmed that the genomic DNA of trypanosomes in pigeon blood was the same as that of T. evansi. This is the first report of the establishment of T. evansi infection in pigeon nestlings.     

Passive Immunization of Pigeons Against Trichomoniasis

SYNOPSIS Nonimmune homing pigeons Columba livia were infected with the Jones' Barn strain of Trichomonas gallinae and subsequently transfused with plasma from acute or chronically infected pigeons harboring one of 3 different strains of T. gallinae. The transfusions were either a single 2 ml dose given one day after inoculation or three 1 ml doses given 0, 5, and 10 days after inoculation. Plasma from pigeons harboring any of the 3 strains was capable of passively immunizing nonimmune birds. All birds which were immunized with plasma from infected pigeons survived until killed at the end of the test period and no visceral lesions were found on necropsy but trichomonads were present in the oropharynx. All controls (untreated or transfused with normal plasma) died of visceral trichomoniasis.
Immune plasma produced some lysis of trichomonads in vitro, and inhibition of motility and vacuolization occurred in some of the non-lysed organisms. The overall lytic activity in vitro affected less than 10% of the suspended trichomonads.     

Impact of urban environment and host phenotype on the epidemiology of Chlamydiaceae in feral pigeons (Columba livia)

Chlamydiaceae are obligate intracellular Gram-negative bacteria found all over the world and known to cause various forms of disease in animals and humans. Urban pigeons are known to be an important reservoir of Chlamydia psittaci, the agent of human psittacosis. In this study, we examined the influence of pigeon houses used to regulate pigeon populations and of melanin-based coloration on several epidemiological parameters of Chlamydiaceae in 708 urban pigeons in Paris. We also identified species and genotypes of Chlamydiaceae present in Parisian populations. First, our results revealed that pigeons roosting and breeding in pigeon houses were equally infected by Chlamydiaceae as those that did not. Second, we found that dark melanic pigeons excreted more Chlamydiaceae than pale melanic ones. Finally, species and strain diversities were very low: all samples were of C. psittaci genotype B. Nevertheless, two atypical Chlamydiaceae were identified based on 16S rRNA and ompA sequences. Our study thus highlights the importance of considering environmental and host phenotype when investigating the epidemiology of infectious diseases.     

Micro diffusion precipitin tests for enteroviruses and influenza B virus

Middleton, G. K., Jr. (Bowman Gray School of Medicine, Winston-Salem, N.C.), H. G. Cramblett, H. L. Moffet, J. P. Black, and H. Shulenberger. Micro diffusion precipitin tests for enteroviruses and influenza B virus. J. Bacteriol. 87:1171-1176. 1964.-A simple micro precipitin gel diffusion test has been adapted to the study of viral antigens. As far as is known from a review of recent literature, this is the first use of the ECHO viruses in precipitin tests and the first attempt to demonstrate by the gel diffusion technique precipitins in a patient's serum after natural virus infection rather than artificial immunization. The principal value of this technique in virology is the rapid identification or qualitative analysis of viral antigen preparations by use of pooled or specific hyperimmune sera. Virus concentrations of 10(7)tcid(50) per 0.1 ml are required for reliable results, but only 0.015 ml of serum is necessary for each test. Virus-serum precipitin reactions were type-specific except for reciprocal precipitation of ECHO 1 and ECHO 8 by their hyperimmune sera. No viral antigens have been found common to two or more virus types among those tested. Precipitins for viral antigens occur frequently in serum of patients after a viral infection and are readily detected by micro precipitin gel diffusion tests. However, this precipitin test remains at present a tool for virus and antigen identification and offers an approach for research appraisal of host response to infections.     

Probability of Detecting Band-Tailed Pigeons During Call-Broadcast Versus Auditory Surveys

Abstract: Estimates of population trend for the interior subspecies of band-tailed pigeon (Patagioenas fasciata fasciata) are not available because no standardized survey method exists for monitoring the interior subspecies. We evaluated 2 potential band-tailed pigeon survey methods (auditory and call-broadcast surveys) from 2002 to 2004 in 5 mountain ranges in southern Arizona, USA, and in mixed-conifer forest throughout the state. Both auditory and call-broadcast surveys produced low numbers of cooing pigeons detected per survey route (x̄ ≤ 0.67) and had relatively high temporal variance in average number of cooing pigeons detected during replicate surveys (CV ≥ 161%). However, compared to auditory surveys, use of call-broadcast increased 1) the percentage of replicate surveys on which ≥1 cooing pigeon was detected by an average of 16%, and 2) the number of cooing pigeons detected per survey route by an average of 29%, with this difference being greatest during the first 45 minutes of the morning survey period. Moreover, probability of detecting a cooing pigeon was 27% greater during call-broadcast (0.80) versus auditory (0.63) surveys. We found that cooing pigeons were most common in mixed-conifer forest in southern Arizona and density of male pigeons in mixed-conifer forest throughout the state averaged 0.004 (SE = 0.001) pigeons/ha. Our results are the first to show that call-broadcast increases the probability of detecting band-tailed pigeons (or any species of Columbidae) during surveys. Call-broadcast surveys may provide a useful method for monitoring populations of the interior subspecies of band-tailed pigeon in areas where other survey methods are inappropriate.     

First detection and genotyping of human-associated microsporidia in pigeons from urban parks

Microsporidia are ubiquitous opportunistic parasites in nature infecting all animal phyla, and the zoonotic potential of this parasitosis is under discussion. Fecal samples from 124 pigeons from seven parks of Murcia (Spain) were analyzed. Thirty-six of them (29.0%) showed structures compatible with microsporidia spores by staining methods. The DNA isolated from 26 fecal samples (20.9%) of microsporidia-positive pigeons was amplified with specific primers for the four most frequent human microsporidia. Twelve pigeons were positive for only Enterocytozoon bieneusi (9.7%), 5 for Encephalitozoon intestinalis (4%), and one for Encephalitozoon hellem (0.8%). Coinfections were detected in eight additional pigeons: E. bieneusi and E. hellem were detected in six animals (4.8%); E. bieneusi was associated with E. intestinalis in one case (0.8%); and E. hellem and E. intestinalis coexisted in one pigeon. No positive samples for Encephalitozoon cuniculi were detected. The internally transcribed spacer genotype could be completed for one E. hellem-positive pigeon; the result was identical to the genotype A1 previously characterized in an E. hellem Spanish strain of human origin. To our knowledge, this is the first time that human-related microsporidia have been identified in urban park pigeons. Moreover, we can conclude that there is no barrier to microsporidia transmission between park pigeons and humans for E. intestinalis and E. hellem. This study is of environmental and sanitary interest, because children and elderly people constitute the main visitors of parks and they are populations at risk for microsporidiosis. It should also contribute to the better design of appropriate prophylactic measures for populations at risk for opportunistic infections.     

Study of the role of pigeons in the dissemination of Cryptococcus neoformans in nature.

Cryptococcus neoformans was recovered from droppings collected within the first 24 h from pigeons experimentally fed with a dose of 5 X 10(6) cells. The fungus proved to multiply well though differently in the sterilized pigeon and chicken excreta seeded with the organism. In both unsterile types of droppings no viable cells of C. neoformans were detected after 4 weeks incubation. Isolated bacterial flora from the intestinal contents of apparently healthy pigeons showed a complete inhibitory effect on the growth of C. neoforms in vitro. It has been concluded that pigeons do not favor multiplication of the fungus in their gut and consequently they do not seem to play an active biological role in dissemination of C. neoformans in nature.     

Attenuated, replication-competent herpes simplex virus type 1 mutant G207: safety evaluation in mice

Herpes simplex virus type 1 (HSV-1) mutants that are attenuated for neurovirulence are being used for the treatment of cancer. We have examined the safety of G207, a multimutated replication-competent HSV-1 vector, in mice. BALB/c mice inoculated intracerebrally or intracerebroventricularly with 10(7) PFU of G207 survived for over 20 weeks with no apparent symptoms of disease. In contrast, over 80% of animals inoculated intracerebrally with 1.5 x 10(3) PFU of HSV-1 wild-type strain KOS and 50% of animals inoculated intracerebroventricularly with 10(4) PFU of wild-type strain F died within 10 days. Similarly, after intrahepatic inoculation of G207 (3 x 10(7) PFU) all animals survived for over 10 weeks, whereas no animals survived for even 1 week after inoculation with 10(6) PFU of KOS. After intracerebroventricular inoculation, LacZ expression was initially observed in the cells lining the ventricles and subarachnoid space; expression decreased until almost absent within 5 days postinfection, with no apparent loss of ependymal cells. G207 DNA could be detected by PCR in the brains of mice 8 weeks after intracerebral inoculation; however, no infectious virus could be detected after 2 days. As a model for latent HSV in the brain, we used survivors of an intracerebral inoculation of HSV-1 KOS at the 50% lethal dose. Inoculation of a high dose of G207 at the same stereotactic coordinates did not result in reactivation of detectable infectious virus or symptoms of disease. We conclude that G207 is safe at or above doses that were efficacious in mouse tumor studies.     

流行性出血热病毒沙鼠肾细胞培养灭活疫苗免疫家兔的抗体反应及保护试验

利用自流行性出血热(EHF)病人血液中分离的EHF病毒浙10株,感染长爪沙鼠肾单层细胞,经培养后,按流行性乙型脑炎的生物制品法规要求研制成灭活疫苗。取1ml经肌肉免疫家兔,6天后即可在血液中检出荧光抗体,至第14～21天为最高峰,荧光抗体效价可达320～2560。经活EHF病毒攻击,有明显的保护作用,保护率达90％以上。疫苗经1:10或1:30稀释后免疫家兔,荧光抗体阳转率仍达100％,但抗体滴度明显降低。肌肉接种优于皮下接种。本研究证明,甲醛灭活EHF病毒可破坏血凝素活性,从而影响疫苗血凝抑制抗体的产生,可能也会影响中和病毒的能力。     

Precipitation of animal plasma proteins by puff-adder venom.

1. Plasma and serum samples obtained from various animals never previously exposed to snakes or snake venom were diffused against different concentrations of puff-adder, Bitis arietans, venom using the double immunodiffusion technique. 2. Depending upon venom concentration, two precipitin arcs could be produced in the case of all plasma samples used. No serum samples showed any arcs except pigeon serum, where one precipitin line was observed. 3. By altering the concentration of snake venom between 1% and 10% when immunodiffusing against plasma a change in position of the precipitin lines was observed and also the disappearance of one or both of the two bands at higher concentrations. This indicates that the arcs observed are in all probability due to precipitation of plasma protein fractions. 4. Previous results indicated that one of the two bands observed when diffusing venom against plasma was due to the precipitation of fibrinogen. By diffusing snake venom against heparin we have now shown that the second band involves this molecule and is not due to another coagulation factor as was suggested previously.