Coy D. Smith

The demonstration of growth stimulating substances forHistoplasma capsulatum andBlastomyces dermatitidis in infusions of starling (Sturnis vulgaris) manure

Comparisons of growth and viability ofH. capsulatum on Sabourauds medium with agar medium plus infusions of soil with and without starling (S. vulgaris) manure indicate that: 1. The largest number of viable conidia and smallest number of hyphal elements were developed on agar plus infusion of loam soil enriched with starling manure (Medium No. 3). 2. The largest volume of growth occurred on the Sabourauds medium which was chiefly vegetative mycelium showing the lowest viability of any of the media tested. 3. Preliminary results indicate thatH. capsulatum will grow as well and produce as many viable conidia on dilute starling manure medium alone as when this medium is mixed or diluted with soil infusion. 4. Repeated passage ofH. capsulatum on infusion of naturally infected soil medium (No. 1) increased the number of total particles, the percentage of microconidia and the total viability. Similar comparisons with soil or starling manure media were made on the growth ofB. dermatitidis except that Sabourauds medium was not included. These comparisons indicate that: 1. The largest number of viable conidia with a small number of hyphal elements was developed on starling manure infusion (medium No. 4). 2. Repeated passage ofB. dermatitidis on starling manure infusion (medium No. 4) resulted in an increase in the number of total particles, the percentage of microconidia, and the total viability.

The susceptibility of Blastomyces dermatitidis to drying.

Summary The effect of drying on the loss of viability of Blastomyces dermatitidis in sterile soil is reported. 1. An average of 90% loss of viability occurred after 5 days storage at room temperature in cotton plugged tubes or in a desiccator. The loss increased to 95% after 11 days or longer. Aliquots of this soil mixed and stored for 10 days in a humidity chamber at room temperature showed a 2-fold increase in viable particles. 2. The loss of viability from storage of soil in tubes after 3 days was 75% with an increase to a 90% loss by 5 days. A 95% loss after 25 days when stored at room temperature or in a desiccator was observed. When soil was placed in a humidity chamber and stored at 4°C for 25 days, only 14% loss in viability occurred. 3. Although B. dermatitidis in soil may lose 90-95% viability after 5-10 days at room temperature, longer storage of up to 90 days does not affect the remaining 5% viable particles. Little change was noted in soil suspended in saline at 4°C after 7 days, but after 30 days, a 95% loss of viable particles was detected. 4. The degree of loss of viability appeared to be correlated with water loss by evaporation.

Comparison of infection of laboratory animals with Blastomyces dermatitidis using different routes of inoculation

Groups of guinea pigs, hamsters and mice of 10 animals each, were injected with 9400, 940 and 94 viable particles of the mycelial phase of Blastomyces dermatitidis. Guinea pigs and hamsters were inoculated intratesticularly, and mice intravenously. It was possible to isolate the pathogen from the 3 species of animals with all the above mentioned doses.Hamsters inoculated intraperitoneally with the same inocula were considerably less susceptible than hamsters and guinea pigs injected intratesticularly. The mice injected intravenously also appeared more susceptible than hamsters inoculated intraperitoneally but due to small numbers of animals used, the assumption is not statistically significant. It appears that the intratesticular route in hamsters or guinea pigs may be a useful tool in the search of B. dermatitidis in its natural environment.

Experimental histoplasmosis and blastomycosis in young pups

Young pups exposed to soil seeded with Histoplasma capsulatum and Blastomyces dematitidis were very susceptible to infection. Of those exposed to H. capsulatum, 88·5% dying 3 weeks or more after contact, yielded positive cultures from various tissues. In compariosn, 57% positive cultures were obtained from the pups dying 3 weeks or more after exposure to Blastomyces dermatitidis. The accumulated percentage of positive cultures obtained from all pups dying in the H. capsulatum group was 75·0% compared to 48·9% in the B. dermatitidis group. After 17 weeks about 95% of the pups died that were exposed to H. capsulatum compared to about 76% at this time exposed to B. dermatitidis.

A comparison of the relative suceptibilities of laboratory animals to infection with the mycelial phase of Blastomyces dermatitidis

Rabbits, guinea pigs, rats, hamsters, and mice were injected intraperitoneally with equal doses of B. dermatitidis prepared from cultures containing large numbers of microspores. Mice and hamsters proved to be the most susceptible to infection.Minimal infective dosages for mice appear to be in excess of 5,000 viable particles. Approximately 66% of the hamsters were infected when inoculated with 2750–4250 viable particles. Optimal incubation time for hamsters appears to be about 6 weeks while maximal recovery from mice was obtained 7–10 weeks after inoculation.A slight variation was shown in the pathogenicity of 4 different isolates of B. dermatitidis inoculated i/p into mice and hamsters, the 2 isolates from human sources tending to be more pathogenic than the 2 canine isolates.

Fifteen isolations of Gymnoascus demonbreunii from canine and soil sources

Eleven isolations of Gymnoascus demonbreunii were obtained from a total of 265 dogs which were killed and tissues cultured for Histoplasma capsulatum. Seven of these isolates were obtained from lung tissue and 4 from mediastinal lymph nodes. The occurrence of this fungus in dogs is thought to be incidental because there were no lesions present nor fungal multiplication in the tissues since the highest number of colonies obtained from one organ was 9 and in the majority of cases only 1. H. capsulatum was isolated in addition to G. demonbreunii from 4 of the above 11 animals.Four isolates of G. demonbreunii were obtained from soils collected from chicken houses where H. capsulatum has also been isolated.After numerous studies, no relationship could be shown between G. demonbreunii and H. capsulatum with the exception of perhaps an environmental one.

Isolation of Histoplasma capsulatum from Soil by Direct Culture Methods.

Summary One hundred and two isolations of H. capsulatum were obtained from 2 soil specimens collected from a starling roosting site by direct culture methods. These isolations were obtained on a yeast extract medium containing penicillin, streptomycin, polymyxin B, and cycloheximide. The number of viable particles of H. capsulatum was estimated per gram of soil to be 4, 500 for soil #1, and 6,300 for soil #2. These numbers were calculated on the basis of the number of colonies isolated from a 1:1000 dilution of the soil specimens.

Distribution of Blastomyces dermatitidis in dogs with skin test and serologic results following airborne infection

Reported is a short term (16 week) experiment to determine the distribution of B. dermatitidis in dogs when infected by the natural inhalation route by receiving about 250,000 viable particles during a 24h. exposure. Four dogs were positive to blastomycin and 1 to histoplasmin prior to exposure. Post exposure, all dogs developed positive skin tests to blastomycin by the 4th week with considerable cross reaction to histoplasmin. Serologic tests showed more poritive CF tests to Histoplasma than to Blastomyces so that the test appears of no practical value. Although 22 tissues were cultured on each dog, only 1 or 7 dogs was positive for B. dermatitidis by 2 weeks. At 4 weeks, 57% of the dogs were positive by culture and at 12, 15, and 16 weeks, all were positive. One dog developed eye lesions and became emaciated.

Supportive evidence by field testing and laboratory experiment for a new hypothesis of the ecology and pathogenicity of canine blastomycosis

This paper offers evidence showing that the pathogenicity of Blastomyces dermatitidis involves the following: 1. Infection with B. dermatitidis in dogs is very severe. Among 33 infected dogs almost 20% died, all but one with eye lesions. In addition, 70% developed skin lesions in the first 66 weeks of observation. 2. The skin test appears to be a useful tool in defining dogs probably infected with B. dermatitidis. 3. Serologic tests do not appear useful as indicators of infection because cross reactions with histoplasmin are frequent. 4. Field skin testing of dogs with blastomycin and histoplasmin indicates that it is extremely useful in detecting infected dog packs and premises. Almost half of more than 100 dogs in the area of the only known blastomycosis epidemic in North Carolina showed specific blastomycin tests as contrasted with an 11–18% rate in mongrel dogs in Kentucky. Skin testing of 20 fox hound packs in Kentucky has revealed 8 premises where 35% or more of the dogs show positive specific blast...

Susceptibility of laboratory animals to Blastomyces dermatitidis using different routes of inoculation.

Experiments have been made to determine the most susceptible animal and best route of inoculation, for the recovery ofBlastomyces dermatitidis. Statistically significant differences were found in the recovery rate ofB. dermatitidis from hamsters and guinea-pigs injected intratesticularly compared with hamsters inoculated intraperitoneally. The intratesticular route in hamsters also appeared significantly more sensitive than the intravenous route in mice. Through both methods it was also possible to recoverB. dermatitidis from some animals inoculated with a suspension of septic garden soil and using only 137 viable particles. The same suspension processed by the oil flotation method with later injection of the supernatant oil into hamsters intratesticularly, permitted the isolation of the agent from all the animals. The processing of soils through this method with later intratesticular inoculation of hamsters should be a useful tool in the search ofB. dermatitidis in nature. Se describen experiencias tendientes a determinar el animal de laboratorio y la mejor ruta de inoculación, para el aislamiento deBlastomyces dermatitidis, a partir de cultivos sobre suelo esterilizado. Se encontró una diferencia significativa en la recuperación del hongo de hamsters y cobayos inoculados por via intratesticular en relación a hamsters inoculados en el peritoneo. La via intratesticular en el hamster se mostró también más sensible que la via intravenosa en el ratón. Mediante ambos méthodos fue posible asimismo recuperarB. dermatitidis de algunos animales inoculados con una suspensión de suelo de jardin no esterilizado, artificialmente contaminado con el hongo e inyectándose aproximadamente 137 partículas viables. El procesamiento del mismo inóculo por el método de flotación en parafina liquida con posterior inyección de la parafina sobrenadante en el hamster por via intratesticular, permitió aislar el agente de todos lo animales inoculados. Se concluye que el procesamiento de suelos por este método, puede ser de gran utilidad para la búsqueda deB. dermatitidis en la naturaleza.