Exposure of rabbits to spores of Aspergillus fumigatus or Penicillium sp: survival of fungi and microscopic changes in the respiratory and gastrointestinal tracts.

Rabbits were exposed to spores of Aspergillus fumigatus by 1 of 2 routes: exposure to aerosols of dry spores or introduction of liquid suspensions of spores directly into the stomach. Rabbits also were exposed to aerosols containing spores of a Penicillium sp. Cultural and microscopic examinations of tissues from the respiratory and gastrointestinal tracts indicated fungi were distributed throughout the gastrointestinal tract of the rabbits within 1 hour after exposure to aerosols of A fumigatus or Penicillum spores. Viable A fumigatus and Penicillium were detected in lung tissues of rabbits for 2 or 3 weeks after inhalation of spores. Aspergillus fumigatus was isolated from the gastrointestinal tract no more than 1 week after aerosol exposure, and Penicillium, not beyond 48 hours. However, when large numbers of A fumigatus spores were introduced directly into the stomach, fungi were isolated from tissues for as long as 16 days after exposure even though the intestinal contents were negative 4 to 7 days after introduction of spores. Tests for precipitating antibody were negative, with one exception, among 26 rabbits surviving for 2 weeks or more. Microscopic changes were more pronounced in rabbits exposed to spores of A fumigatus than in rabbits exposed to Penicillium spores.     

Rapid hematogenous dissemination of Aspergillus fumigatus and A. flavus spores in turkey poults following aerosol exposure

Cultures of blood samples taken from 3-week-old turkey poults exposed to aerosolized spores of either Aspergillus fumigatus or A. flavus for 15 min yielded organisms. Aspergillus fumigatus was also isolated from brain and liver tissue samples taken immediately after exposure. Exfoliated cells from the lungs of 10 poults exposed to aerosolized spores of A. fumigatus were allowed to attach to glass slides in tissue culture. Several types of the fixed and stained cells had attached or ingested spores of A. fumigatus. Macrophages were the predominant cell type with ingested spores, although other cell types may be involved in the transport of spores of A. fumigatus into the blood stream after aerosol exposure.     

A case of aspergillosis in a broiler breeder flock

A case of aspergillosis in a broiler breeder flock having respiratory and nervous system problems caused by Aspergillus fumigatus and Aspergillus niger is documented. Dyspnea, hyperpnea, blindness, torticollis, lack of equilibrium, and stunting were observed clinically. On postmortem examination of the affected birds, white to yellow caseous nodules were observed on lungs, thoracic air sacs, eyes, and cerebellum. Histopathologic examination of lungs and cerebellum revealed classic granulomatous inflammation and cerebellar lesions, necrotic meningoencephalitis, respectively. No lesions were noted in the cerebrum histopathologically. Aspergillus hyphae were observed in stained sections prepared from lesioned organs. Fungal spores and branched septate hyphae were observed in direct microscopy. Aspergillus fumigatus and A. niger were isolated from the inoculations prepared from the suspensions of organs showing lesions.     

Pathogenesis of Aspergillus fumigatus infection in pigeons in the Sudan.

The pathogenesis of a pigeon isolate of Aspergillus fumigatus in a local breed of pigeons in the Sudan was tested. The spores inoculated intravenously resulted in an acute disease with 100% of mortality within six days. At necropsy, pinpoint and miliary lesions were prominent in the liver, spleen, lungs and kidneys. Histopathological examination detected lesions in the liver, heart, lungs, kidneys, spleen and brain. Hyphae and/or spores were encountered in all these organs. The presence of A. fumigatus was confirmed by reisolation from the liver, lungs and kidneys.     

Development of a method for the identification, using the polymerase chain reaction, of Aspergillus fumigatus isolated from ostriches

Objective To develop a method for identifying DNA of Aspergillus fumigatus from ostriches, using the polymerase chain reaction (PCR). A fumigatus is the principal causative agent of avian aspergillosis.
Design A biochemical trial.
Sample population Twelve Aspergillus fumigatus isolates and three other Aspergillus species.
Procedure PCR primers that were based on the sequence of the alkaline protease gene from human isolates of A fumigatus were used.
Results We successfully tested the method on ostrich isolates from five states and showed that the test is specific for A fumigatus.
Conclusions In most cases the DNA sequence of A fumigatus isolates from ostriches is similar to that of human isolates. DNA sequences vary significantly among A fumigatus isolates, including those from affected ostriches in the same flock. The genetic variation may be used to trace aspergillus infections in ostrich flocks and determine if the disease is transmitted by contact with infected birds.     

Histopathologic and antibody responses of rabbits exposed to aerosols containing spores of Aspergillus fumigatus: comparison of single and multiple exposures

Resistance to pulmonary aspergillosis was studied in groups of rabbits exposed to aerosolized spores of Aspergillus fumigatus for 15 minutes on successive days for a total of 10, 7, or 4 exposures or a single exposure. The results of the study demonstrated that exposure of rabbits to spores for 15 minutes on 10 successive days did not result in an accumulation of viable spores in excess of those present in the lungs of rabbits exposed a single time. The tissue response in the lungs of the rabbits exposed at multiple times was more intense than that in the rabbits exposed once, but resolution of the lesions occurred similarly in terms of time and completeness of resolution. The duration of the antibody response as determined by a passive hemagglutination test and by enzyme-linked immunosorbent assay correlated with the number of exposures to spores, in that rabbits exposed 10 or 7 times to aerosolized spores remained positive longer than did those exposed fewer times. The results of the precipitin tests in agar gel were negative in all the rabbits but one.     

Antifungal activity of enilconazole on experimental Aspergillosis in chickens

Day-old chickens were infected with various inoculum sizes of airborne spores of Aspergillus fumigatus, which caused high rates of mortality and morbidity. Fumigation with enilconazole smoke pellets was highly successful in preventing mortality, in reducing morbidity, and in neutralizing growth inhibition of infected chickens. No side effects were observed.     

The antifungal activity of natamycin toward molds isolated from commercially manufactured poultry feed

The antifungal activity of natamycin, a polyene antifungal compound, was evaluated on molds isolated from commercial poultry feed. The antifungal activity was measured by determination of the minimal inhibitory concentration (MIC) for natamycin on molds growing on semisolid microbiological medium (potato dextrose agar) containing pure natamycin at concentrations ranging from 0 to 200 mg/liter. Natamycin exhibited a high degree of antifungal activity against the 191 isolates of aspergilli used in this study, with average MIC values ranging from 5.08 to 40.1 mg/liter for Aspergillus fumigatus and Aspergillus parasiticus, respectively. Natamycin was also equally effective in inhibiting the growth of nonaflatoxigenic compared with aflatoxigenic isolates of Aspergillus flavus and A. parasiticus. Natamycin was also efficacious against molds other than aspergilli, with MIC values ranging from 2.15 to 5.80 mg/liter for Paecilomyces and Rhizopus spp., respectively. Natamycin exhibited apparent sporicidal activity against spores of toxigenic strains of Fusarium moniliforme and A. parasiticus but not Penicillium rubrum. This sporicidal activity was evident only when spores were exposed to an in vitro concentration of natamycin of 25 mg/liter or higher for a period of time of at least 12 hr. The growth inhibiting activity of natamycin was more pronounced compared with the sporicidal activity.     

Mass occurrence of mycotic tracheitis in chickens

An enzootic of chick mycosis, caused by the spores of the fungus Aspergillus fumigatus, is described. The mycotic infection affected the respiratory tract of the birds; pathological changes were located mainly in the region of the trachea. The changes had the nature of diphtheroid necrotic inflammation destroying the mucous membrane and causing almost an obstruction of the trachea. Deposits of granulomatous inflammation, containing fungus elements, were detected in the peritracheal tissue, and in individual birds also in the lungs. Litter contaminated with Aspergillus was the source of infection.     

Fungal isolation and identification in 21 cases of guttural pouch mycosis in horses (1998-2002)

This aetiological study of guttural pouch mycosis (GPM) in the horse was based on the retrospective study of 21 horses brought into the National Veterinary School of Lyon (France) between 1998 and 2002. Biopsies were taken from the lesions caused by GPM during endoscopic examination. In 87% of the cases, direct examination gave positive results, whereas 43% of the cultures were found to be negative. The main fungi observed were Aspergillus fumigatus (in three cases), A. versicolor (in two cases, together with other fungi), and A. nidulans and A. niger (one case each). In six cases, the Aspergillus species could not be identified. In two cases, cleistothecia and/or Hulle cells were observed. In three cases, fungi other than Aspergillus were seen, mixed or not with Aspergillus. These results underline the importance of Aspergillus fumigatus in the development of GPM in horses.     

Pathological, clinical and mycological findings in experimental aspergillosis infections of starlings

A group of 24 wild starlings (Sturnus vulgaris), in undefined age categories but at least post-pubertal, constituted the material of this study. Six starlings were kept as a control group and 18 starlings served as the infection group. The starlings in the infection group were infected with inoculums of 1.35 x 10(6)/0.2 ml Aspergillus fumigatus via intratracheal route whereas the control group was administered only placebo in the same way. Six, four, two, four and two birds died on 2, 3, 4, 5 and 6 days post inoculation respectively. At the necropsy of the dead birds, caseous foci were determined in the lungs, on the air sacks, myocardium, thoracic wall and abdominal serosa. In the histopathological examination of the white-yellowish caseous foci ranging from pinhead to chick pean in size, necrotic granulomatous foci consisting of macrophages, heterophil leukocytes and gigant cells encapsulated with a fibrous tissue were observed. Hyphae and spores of A. fumigatus were determined in these foci using the Gridley staining method.     

Aspergillus fumigatus infection in an ostrich (Struthio camelus)

An 11-month-old female ostrich (Struthio camelus) had become gradually emaciated over a 2-week period and subsequently died. Necropsy revealed white to green mold growth on the walls of caseous thickened air sac membranes and multiple white necrotic foci in the lungs and liver. Histologically, the multiple exudative, necrotic and granulomatous lesions were compatible with mycotic infection in the air sacs and lungs, and hyphae positively reacted with a monoclonal antibody (Mab-WF-AF-1) to Aspergillus fumigatus wall fractions. Multifocal hepatic necrosis was also found, and several spores were observed in the blood vessels. Fungal culture of these lesions yielded pure growth of A. fumigatus. This is an established case of fatal A. fumigatus infection in an ostrich reared in Japan.     

IgE-bearing cells in bronchoalveolar lavage fluid and allergen-specific IgE levels in sera from RAO-affected horses

Recurrent airway obstruction (RAO) is a common condition in stabled horses characterized by small airway inflammation, airway neutrophilia and obstruction following exposure of susceptible horses to mouldy hay and straw and is thus regarded as a hypersensitivity reaction to mould spores. However, the role of immunoglobulin E antibodies (IgE) in the pathogenesis of RAO is unclear. We hypothesized that the number of cells with receptor-bound IgE in bronchoalveolar lavage fluid (BALF) and IgE levels in serum would be higher in RAO-affected than in healthy horses living in the same environment. Therefore, IgE-positive (+) cells were identified by immunocytochemistry on cytospins from BALF and counted. IgE levels against the mould extracts Aspergillus fumigatus (Asp. f.) and Alternaria alternata (Alt. a.) and the recombinant mould allergen Aspergillus fumigatus 8 (rAsp f 8) were measured by enzyme-linked immunosorbent assay (ELISA) in the sera of seven RAO-affected and 22 clinically healthy mature horses housed in the same conventional stable environment. After correcting for the number of neutrophils, there were no significant differences in IgE+ cells on cytospins from BALF between both groups of horses (5% versus 7%, P > 0.1). Serum IgE levels against the mould extracts were significantly higher in RAO-affected than in clinically healthy horses [median = 119 versus 66 relative ELISA units (REU), P < 0.05]. Furthermore, significantly more RAO-affected than healthy horses had detectable serum IgE against the recombinant allergen rAsp f 8 (4/7 and 3/22, respectively, P < 0.05). Age had no significant effect on BALF cell ratios or on specific serum IgE levels. These results show that high IgE levels against mould antigens are associated with RAO under controlled environmental conditions but ranges of mould-specific serum IgE levels overlapped too much between diseased and clinically healthy animals to be of any diagnostic value. Further studies are needed to assess whether IgE-mediated reactions contribute to the pathogenesis of RAO.     

Aspergillus fumigatus infection in two wild Eurasian black vultures (Aegypius monachus Linnaeus) with carbofuran insecticide poisoning: a case report

Aspergillus spp. are opportunistic pathogens which cause pulmonary aspergillosis in animals and humans with compromised immune systems. Two Eurasian black vultures (Aegypius monachus Linnaeus) were found dead or clinically ill from carbofuran insecticide during the winter of 2004. Carbofuran was detected in the stomach contents by gas chromatograph-mass spectrometry. Gross lesions showed severe granulomatous pneumonia and serofibrinous pleuropneumonia in both birds, with most lesions restricted to the pulmonary system. Histological lesions included pyogranulomatous pneumonia and suppurative parabronchiolitis/pleuritis/air sacculitis with a number of septated fungal hyphae, suggesting severe pulmonary aspergillosis. Fungal isolates from each vulture were identified as Aspergillus fumigatus by both lactophenol cotton blue staining and genetic analysis. This is the first report of pulmonary aspergillosis caused by A. fumigatus in wild Eurasian black vultures and suggests that Aspergillus infection could be an important cause of death in these birds which migrate from Mongolia to Korea during the winter. The incidence of the disease may be related to impaired immunity caused directly or indirectly by carbofuran poisoning.     

Effects of some egg characteristics on the mass loss and hatchability of ostrich (Struthio camelus) eggs

1. This study was conducted to examine some egg characteristics and determine the effects of eggshell thickness and eggshell porosity on water loss and hatchability of eggs in ostriches. 2. Shell thickness did not correlate significantly with hatchability. However, eggs of low shell thickness lost more mass (13.03%) than those with intermediate (11.22%) and high (10.36%) shell thickness. Mass loss during incubation was higher in hatched (11.98%) than unhatched eggs (11.09%). Shell thickness was negatively correlated to egg mass loss (r = -0.65). 3. The pore density was correlated with hatchability. Hatchability was 50% lower in eggs with low pore densities (40.93%) than with high densities (80.94%). Pore density was positively correlated with egg mass loss (r = 0.63). Incubation mass losses of hatched and unhatched eggs were not significantly different. 4. Mean eggshell water vapour conductance (G) value and shell conductance constant (k) were 87.77 +/- 4.21 mg H2O/d/Torr and 2.44 respectively (n = 15). 5. Because of eggshell functional properties and resulting low egg mass loss hatchability is low when ostrich eggs are artificially incubated. The mass of eggs used in the experiment was relatively high and their eggshell water vapour conductance was low. As a result, egg incubation mass loss was lower than it should be. It is concluded that incubator humidity should be low (25%) to allow enough mass loss during incubation from the eggs.     

Moisture content and moisture quantity of sweated chicken eggs in 2 storage environments

There are instances where shell eggs may be moved from refrigeration into ambient temperature with high humidity, such as before wash and during transportation. Under these conditions, it is of concern that bacteria on wet eggs can grow and migrate through the shell pores into the egg. Objectives of this experiment were: 1) to compare 3 methods of quantifying condensate on eggs and 2) to quantify condensate on refrigerated shell eggs at 2 temperatures (22°C and 32°C). For objective 1, 270 fresh shell eggs (3 replications, 90 eggs per replication) were stored at 4°C, 60% relative humidity (RH), then placed at 22°C, 60% RH for 1 h. After this time, 30 pre-weighed eggs were randomly selected and weighed. Thirty eggs were thoroughly wiped with pre-weighed paper towels to collect condensate. Thirty eggs were evaluated with a pinless moisture meter for quantifying egg condensate, which was found to be an ineffective method. There was no difference in quantifying egg condensation by egg weight or weight of moisture absorbed on a paper towel (0.2% vs. 0.19% percentage gain mL condensation/egg surface area) (P > 0.05). For objective 2, 104 fresh eggs formed condensation at 2 temperatures (22°C and 32°C, 60% RH). Each egg weight was continuously recorded from the beginning of condensation formation to the point where the egg reached a constant weight. There was a difference found in the time it took for an egg to reach maximum condensation (11 min at 32°C, 17 min at 22°C), as well as completely dry (25 min at 32°C, 34 min at 22°C) between the 2 temperatures (P < 0.05).     

Bacterial contamination of eggs and behaviour of poultry flocks in the free range environment

The free range production system is becoming more common in Australia and is expected to increase. Free range hens are exposed to more stressors in comparison to hens from barn and cage systems and it is suggested that stress can increase bacterial shedding on eggs. The aims of this study were to examine the level of total bacteria and Enterobacteriaceae populations, as well as the presence of Salmonella and Campylobacter, in eggs collected from two free range flocks on two different farms and to conduct longitudinal observations of the behaviour and welfare of hens in the free range production system. Hen age (weeks) was shown to have a significant effect (increase) on the level of total bacteria on the egg shell surface and in shell pores, as well as having an effect on feather condition score. As the hens aged, the frequency of external visual egg characteristics increased, as did feather condition score (where feather condition was poorer). These observations indicate areas which should be investigated further to improve the food safety of eggs and optimise the welfare of free range hens.     

Vertical transmission of Ovipleistophora ovariae (microspora) within the eggs of the golden shiner

Fertilized eggs collected from broodfish infected by Ovipleistophora ovariae were tested by quantitative polymerase chain reaction (PCR) and found to be positive for the O. ovariae genome at 7.77 X 10(2) to 3.26 x 10(7) copies per microgram of host DNA. Fry hatched from these eggs contained from 1.37 X 10(2) to 9.89 X 10(6) copies of the O. ovariae genome per microgram of host DNA. Surface treatments of fertilized eggs with 150 mg formalin/L (used by farms as a fungicide) or a 1.5% solution of sodium sulfite (which removes the adhesive egg matrix) did not reduce vertical transmission to fry. Treatment of eggs with a 10% solution of bleach or a proprietary commercial DNA denaturant did not reduce the number of egg-associated copies of the O. ovariae genome. Histology of ovaries of infected fish demonstrated spores within the oocytes. However, no spores were observed by histology in positive fry hatched from infected eggs. The PCR and histological demonstration of the presence of O. ovariae spores in oocytes and fry, and the failure of strong DNA denaturants to reduce egg-associated copies, give evidence that O. ovariae is vertically transmitted within eggs.     

Experimental Aspergillus fumigatus infection in quails and results of treatment with itraconazole

This study was performed to investigate (i). the clinical, histopathological and biochemical changes in quails (Coturnix coturnix japonica) with experimentally induced aspergillosis; and (ii). the efficiency of itraconazole treatment on these infected birds. A total of 18021-day-old male quails was randomly divided into three groups (control, infected untreated and infected treated), each containing 60. The experimental infection was set by intratracheal inoculation of 0.2 ml inoculum of Aspergillus fumigatus (CBS 113.26 strain) consisting of approximately 2.7 x 106 spores/ml. Two days after the inoculation, general clinical signs of aspergillosis in the respiratory tract were observed. In the histopathological examination, caseous foci were found in lungs, trachea and on airsacs. All quails died in the infected untreated group. Aspergillus fumigatus was isolated from the various organs of all dead quails. There was no significant change in serum aspartate aminotransferase (AST) and gamma-glutamyltransferase (GGT) activities in infected untreated birds compared with controls. However, alanine aminotransferase (ALT) activity, albumin and calcium levels, and albumin/globulin (A/G) ratio were lower while phosphorus and globulin levels were higher in the infected untreated group than in controls. Each quail in the infected treated group was given 10 mg/kg/day itraconazole via drinking water for 7 days immediately after the first clinical findings. Although all quails died in the infected untreated group, 41 quails survived in the itraconazole treatment group. Biochemical values also returned approximately to the control levels after the treatment. The conclusion was drawn that aspergillosis in the quails might cause economical losses because of high mortality. Oral itraconazole treatment of aspergillosis might lower the mortality rate in quails.     

Omphalitis associated with Aspergillus fumigatus in poults

Omphalitis associated with aspergillosis was diagnosed in four cases of commercial turkey poults ranging in age from 3 to 9 days old. In two cases, the mycotic agent present in the yolk sac was isolated and identified as Aspergillus fumigatus. In the other two cases, the fungi were identified as Aspergillus sp. on the basis of morphologic characteristics of the fungi in tissue sections. The fungi present were further confirmed to be of the genus Aspergillus by immunohistochemistry. Omphalitis by A. fumigatus infection has not been documented before.