Sequential pathological studies in the udder of goats intramammarily infected withAspergillus fumigatus

Intramammary inoculation of goats withAspergillus fumigatus spores resulted in the development of mastitis with characteristic gross and microscopic lesions. The mastitis and the lesions were restricted to the infected udder halves only and there was no dissemination of infection to other tissues of the body. The experiment was continued for 45 days. Gross changes in the infected udder were observed up to the 45th day post-infection. The lesions, in general, included variable sized abscesses in the first 15 days followed by development of varying sized greyish-white nodules in the infected udders. Microscopic changes consisted of granulomatous reaction with well developed granulomas in the infected udders. Hyphae and spores ofAspergillus fumigatus could be demonstrated in sections of the infected udders up to 45 days after infection. Reisolation of the fungus consistently was achieved up to 45 days. It is concluded that intramammary inoculation ofAspergillus fumigatus spores in goats leads to chronic granulomatous mastitis.     

Humoral immunoresponse of pigeons toAspergillus fumigatus antigens

The aim of this study was to develop an immunological model of avian Aspergillosis by studying the humoral response of pigeons toAspergillus fumigatus antigens. Immunization was performed by administering weekly injections ofA. fumigatus extracts for 70 days (10 weeks). A new booster injection was given 270 days (9 months) following the last immunization. Results showed an earlyAspergillus-specific humoral immunoresponse which reached a maximum level at 42–63 days (6–9 weeks) post-immunization. Using the ELISA method, it could be observed thatA. fumigatus-specific IgG became elevated in the 2nd week and reached a maximum titre at 63rd day (9th week). In contrast,A. fumigatus-specific IgM levels appeared early showing maximum levels at the 2nd week, after which they declined despite the maintenance of antigenic stimulation. Termination of immunization resulted in the decrease of specific humoral immunoresponse with minimal levels of specific antibodies detectable 210 days (7 months) later. A booster injection given at 270 days (9 months) induced a very fastAspergillus-specific IgM and IgG immunoresponse, reaching levels of antibodies similar to those observed during the immunization period.     

Experimental aspergillosis in Japanese quails (Coturnix coturnix japonica)

Intratracheal inoculation of 2-week-old quail chicks with Aspergillus fumigatus resulted in the development of clinical signs within 24 h of infection. These were characterized by anorexia, depression, accelerated respiration and gasping followed by death. The acute course of the disease lasted for 7–10 days followed by recovery in the surviving chicks. The overall mortality during a 6-week observation period was 20%.Although the mean body weight of A. fumigatus infected quail chicks continued to be slightly lower throughout the experiment but the difference, in comparison to controls, was not significant except at 42 days post-infection.There was no appreciable difference in the mean values of Hb, TEC, PCV, MCV, MCH and MCHC between the infected and control chicks at any stage of infection but TLC revealed a leucocytosis from 2–7 days which was the result of increase in the relative percentage of heterophils and decrease in lymphocytes.     

Biochemical and Immunological Studies on Aspergillus

Two kinds of polysaccharides, glucan and glycopeptide (galactomannan-peptide), were obtained from Aspergillus fumigatus (IFO 5840) by extraction with 50% pyridine and were purified by fractional precipitation with acetone, by a column chromatography on Dowex-50 and DEAE-cellulose and by the gel filtration on Sephadex G-50 and G-200. The glycopeptide, designated APSK-66 fraction, showed both an Arthus and delayed type (tuberculin type) skin reactions in sensitized rabbits and guinea pigs. By treatment with proteolytic enzymes, the delayed type skin reactivity of APSK-66 fraction was reduced but the Arthus type skin reactivity was not affected. However, the Arthus type skin reactivity of APSK-66 fraction was completely lost by periodate oxidation, and the delayed type skin reactivity of APSK-66 fraction was retained. The APSK-66 fraction showed precipitation, complement fixation and passive hemagglutination reactions with rabbit antisera against A. fumigatus. Glucan, designated as APSK-33 fraction, did not show any immunological activity when tested in the present experiment. The chemical structures of the glucan and galactomannan were discussed.     

Sequential pathological studies in goats infected intratracheally withAspergillus fumigatus

Intratracheal inoculation of goats withAspergillus fumigatus spores resulted in the development of characteristic gross and microscopic lesions. The lesions were restricted to lungs and there was no dissemination of infection to other tissues of the body except liver in one goat 16 days after infection. The experiment was continued for 37 days. Gross changes in lungs were observed up to the 24th day post-infection. The lesions, in general, included congestion and oedema in the first 6 days followed by the development of varying greyish-white nodules in the lungs. Microscopic changes consisted of granulomatous reaction with well developed granulomas in lungs. Hyphae and conidiophores with fruiting bodies ofAspergillus fumigatus could be demonstrated in sections up to 24 days of infection. Reisolation of the fungus consistently was achieved up to 24 days. It is concluded that intratracheal inoculation ofAspergillus fumigatus spores in goats leads to pulmonary aspergillosis up to 24 days.     

A study of the mode of transmission of maize streak virus by Cicadulina mbila using an enzyme-linked immunosorbent assay

Two batches of Cicadulina mbila were given two distinct acquisition access periods (AAP) (3 h and 50 h) on maize plants infected with maize streak virus (MSV). Infectivity assays on susceptible maize were carried out 1, 3, 10, 17, 26 and 35 days after the AAP. Transmission efficiency was significantly higher for C. mbila subjected to the 50-h AAP. At the same time as the infectivity assays, the amount of MSV in each leafhopper was determined by an indirect double antibody sandwich (IDAS) ELISA. There were more ELISA-positive insects after the 50-h AAP than after the 3-h AAP. In the group given a 3-h AAP, only 7% of the insects tested between day 1 and 35 were found to be positive by ELISA. In contrast, after the 50-h AAP, the majority of C. mbila were positive, yet a decrease in ELISA-positive insects was noticed from day 17 onwards. Using a calibration curve obtained with purified virus, as little as 0.15 ng of MSV per insect could be measured by the IDAS-ELISA. A mean value of 0.36 ng of MSV per C. mbila was found 3 days after the 50–h acquisition, whereas 14 days later there was only 0.20 ng of virus per insect. For comparison, when leafhoppers were kept on infected maize, they displayed substantial accumulation of MSV up to an average of 3.83 ng of MSV per insect after 35 days of continuous acquisition. The amount of virus per insect detected in females was usually greater than the amount detected in males. Our results suggest that MSV does not multiply in C. mbila and contribute to the understanding of the persistence of transmission efficiency in the absence of virus multiplication.     

Bronchoalveolar lavage in an immunosuppressed rabbit model of invasive pulmonary aspergillosis

A rabbit model of invasive aspergillosis has been used to investigate the pathogenesis of Aspergillus infection in the immunosuppressed host. The animals received hydrocortisone daily and a single dose of cyclophosphamide 2 days prior to intratracheal instillation of conidia from Aspergillus fumigatus. Bronchoalveolar lavage (BAL) was performed in 3 infected and 2 control saline treated animals sacrificed on days 1, 2, 4, 7 and 10 following inoculation. Infective load within the lung was quantified using an assay for chitin which is an important component of fungal cell walls (in particular the hyphal cell wall) and is not present in vertebrate tissue. The total BAL white cell count did not discriminate between infected and saline treated animals and Aspergillus was cultured from one lavage specimen only. Infected animals developed a marked neutrophil alveolitis by day 2 in contrast to a near total absence of neutrophils in the lavages of the control animals. Phagocytosis of conidia by alveolar macrophages was prominent but did not prevent progressive infection as confirmed by measurement of lung chitin. This pattern of cellular response within the alveolar airspace reflects the complex nature of the response to Aspergillus infection in the immunosuppressed host.     

Experimental Vaccination of Chicks with Plasmodium gallinaceum Sporozoites. I. Circumsporozoite Proteins Are Expressed by Sporozoites Recovered from Both Salivary Glands and Midguts of Mosquitoes1

Immunogenicity of Plasmodium gallinaceum Sporozoites for chicks and their in vitro reactivity with normal and specific immune sera were studied. Two sporozoite populations recovered from experimentally infected Aedes fluviatilis were used: sporozoites from salivary glands and sporozoites from midgut oocysts. Populations seven to nine days old of sporozoites recovered from salivary glands were infective for all chicks until the chicks were three weeks old; however, sporozoites recovered from midguts containing oocysts infected these chicks only if isolated on days 8–9, but not on day 7 after the mosquitoes' infective blood meal. Infectivity of the sporozoites was lost after exposure to ultraviolet (UV) light (30 min) or X-rays (13 krad). Inactivated sporozoites from both sources proved highly immunogenic to chicks that were immunized by several intravenous or intramuscular injections. These parasites elicited a strong humoral immune response in the chicks, as measured by the circumsporozoite precipitation (CSP) reaction. The levels of the CSP antibodies were similar with sporozoites from both sources, there being no detectable differences in the percentage of reactive sporozoites or the intensity of the CSP reaction with sera containing antibodies to either sporozoites from salivary glands or sporozoites from oocysts. These results provide the first evidence that avian malaria sporozoites express the circumsporozoite protein that has been extensively characterized in mammalian malaria (rodent, simian, human sporozoites). Furthermore, we observed that the yields of sporozoites obtained from mosquito midguts, on days 8 and 9 of the P. gallinaceum infection, were at least twice as great as those obtained by salivary gland dissection, even 20 days after a blood meal. This is an advantage since obtaining the midguts is less tedious, as well as more efficient and faster.     

Recent studies on aspergillosis in turkey poults

A review of the studies on aspergillosis in turkey poults at the National Animal Disease Center include limited field studies, pathogenicity studies, and vaccine development. Natural ventilation in turkey rearing houses was effective in reducing airborne propagules of four major fungal genera, but the effectiveness of ventilation appeared to be limited by the width of the building. Aspergillus fumigatus was more effective than A. flavus in producing mortalities in aerosol exposed poults. Toxigenicity of A. flavus did not enhance its pathogenicity, and no apparent aflatoxin production occurred during pathogenesis in infected turkey poults. Spores of A. fumigatus were disseminated quite rapidly in poults exposed to aerosols, and alveolar macrophages from respiratory lavages taken immediately after exposure contained spores of A. fumigatus. Vaccines produced from germlings of A. fumigatus and administered to turkey poults were the most efficacious of five vaccines tested against challenge exposure to aerosols of A. fumigatus spores.     

Chromatographic fractionation of Aspergillus endotoxins

Summary Crude preparations of the endotoxins extracted from the mycelia ofAspergillus fumigatus andAspergillus flavus, after preliminary concentration by ammonium sulfate, have been fractionated by column chromatography on DEAE-cellulose. Although the biological activity of the chromatographed preparations was not limited to a single fraction, examination of the most active fractions by starch gel electrophoresis showed no bands common to the two nephrotoxins.The highest hemolytic and toxic activities of the fumigatus toxin were found in different fractions of the chromatographed material, and starch gel electrophoresis showed no bands common to these two fractions.The molecular weight of the flavus toxin has been estimated to be in the range of 32,000 to 34,000 as judged by the results of ultracentrifugation and microelectrodialysis in starch gels of the most toxic fractions.Both of the toxins have been shown to contain small amounts of hexosamine and larger amounts of non-amino sugars.Presented in part at the 46th Annual Meeting of the American Society of Biological Chemists, Atlantic City, New Jersey, April, 1962.     

Aerobiological,biochemical and immunological studies on some of the dominant <Emphasis Type="Italic">Aspergillus</Emphasis> species of South Assam (India)

Two years atmospheric survey of air-borne Aspergillus was carried out in the environmental conditions of South Assam. The survey revealed a total of 16 different species of Aspergillus with marked seasonal and annual variations. Aspergillus fumigatus was found to be the dominant atmospheric fungal species followed by Aspergillus flavus, Aspergillus niger, etc. Among the sample extracts tested, highest quantity of soluble protein was recorded in Aspergillus fumigatus (95.0 mg/g) whereas highest quantity of soluble carbohydrate (40.8 mg/g) and free amino acid (135.0 mg/g) was recorded in the sample extract of Aspergillus niger per gram of dry weight, respectively. The highest numbers of protein polypeptide bands were detected in the sample extract of Aspergillus fumigatus followed by Aspergillus flavus and lowest in Aspergillus niger. The maximum numbers of immunoglobulin E binding protein fractions were found in Aspergillus fumigatus, followed by Aspergillus flavus, Aspergillus clavatus, etc.     

Harvest and survival of Aspergillus fumigatus fresenius spores

Astract An apparatus used for the collection ofAspergillus fumigatus Fresenius spores from agar cultures is described. Dry spores ofA. fumigatus collected with this apparatus maintained their viability for at least 8 weeks.Mention of products used in this investigation does not constitute endorsement by the USDA.     

Degradation and corrosive activities of fungi in a diesel–mild steel–aqueous system

The fungi Aspergillus fumigatus, Hormoconis resinae and Candida silvicola were isolated from the fuel/water interfacial biomass in diesel storage tanks in Brazil. Their corrosive activities on mild steel ASTM A 283-93-C, used in storage tanks for urban diesel, were evaluated after various times of incubation at 30 °C in a modified Bushnell–Haas mineral medium (without chlorides) with diesel oil as sole source of carbon. Their ability to degrade diesel oil was evaluated after growth for 30 and 60 days. The fungus Aspergillus fumigatus and the consortium of all three organisms showed the highest production of biomass; A. fumigatus gave the greatest value for steel weight loss and produced the greatest reduction in pH of the aqueous phase. Solid phase microextraction (SPME) showed that the main acid present in the aqueous phase after 60 days incubation with A. fumigatus was propionic acid. Polarization curves indicated that microbial activity influenced the anodic process, probably by the production of corrosive metabolites, and that this was particularly important in the case of A. fumigatus. This fungus preferentially degraded aliphatic hydrocarbons of chain lengths C₁₁--C₁₃ in the diesel, producing 47.7, 37.5 and 51% reductions in C₁₁, C₁₂ and C₁₃, respectively. It produced more degradation than the consortium after 60 days incubation. It is likely that the presence of other species in the consortium inhibited the growth of A. fumigatus, thus resulting in a lower rate of diesel fuel degradation.     

Sequential pathological studies in Japanese quails infected experimentally with Aspergillus fumigatus

Intratracheal inoculation of young quail chicks with Aspergillus fumigatus spores resulted in the development of characteristic gross and microscopic lesions. The lesions were restricted to respiratory tract and there was no dissemination of infection to other tissues of the body.Gross changes in lungs and air sacs were observed within 24 hours and continued up to 20 days while in trachea these were noticed from the 3rd to the 9th day post-infection. The lesions, in general, included congestion and focal haemorrhages in the first 2 days followed by the development of varying-sized greyish-white nodules in the lungs, air sacs and trachea.Microscopic changes consisted of congestion, haemorrhages and a diffuse cellular infiltration in the first 2 days followed by granulomatous reaction with well developed granulomas in lungs, air sacs and trachea. Spores and developing hyphae of Aspergillus could be demonstrated in sections from 24 hours to 20 days of infection.Reisolation of the fungus was consistently achieved from the lungs, air sacs and trachea up to 14 days.     

Infection of ‘Candidatus Phytoplasma ulmi’ reduces the protein content and alters the activity of detoxifying enzymes in Amplicephalus curtulus

It has been reported that insecticide‐detoxifying enzymes such as glutathione S‐transferases (GST) and esterases are affected by microbial infections in hemipteran insect vectors. The total protein content, and GST and α‐ and β‐esterase activities were quantified in ‘Candidatus Phytoplasma ulmi’‐infected and uninfected adults of Amplicephalus curtulus Linnavuori & DeLong (Hemiptera: Cicadellidae) at 25, 35, and 45 days after the acquisition access period (AAP) in the head‐thorax and abdomen sections. The total protein content was lower in phytoplasma‐infected leafhoppers 25, 35, and 45 days after the AAP. Thirty‐five days after the AAP, the GST and β‐esterase activities had increased (26 and 69%, respectively) compared to the control. However, 45 days after the AAP, the phytoplasma‐infected leafhoppers displayed lower GST (87%) and β‐esterase (253%) activities than the uninfected individuals. On the other hand, the α‐esterase activity proved to be unaffected by the phytoplasma infection. Forty‐five days after the AAP, females had a higher phytoplasma titer (46%) in their head‐thorax than in their abdomen sections, whereas males showed a higher titer in their abdomens (75%). In addition, the GST and β‐esterase activities in the abdomen were affected negatively by 96–98% as a result of the increasing ‘Ca. Phytoplasma ulmi’ titer. These results indicate that an infection of ‘Ca. Phytoplasma ulmi’ alters the metabolic activities of A. curtulus.     

Natural occurrence of gliotoxin in turkeys infected with Aspergillus fumigatus,Fresenius

Thirteen samples of infected turkey lung tissue from cases of airsacculitis were collected either at the processing plant or from a local turkey farm and subjected to cultural and gliotoxin analysis. Aspergillus fumigatus was isolated from 6 of the 13 samples; all isolates were determined to be gliotoxin producers when grown in laboratory culture and assayed by HPLC procedures. Gliotoxin was isolated from 5 of the 13 tissues but was not isolated from all tissues that were infected with A. fumigatus. Gliotoxin was isolated from two tissues from which no A. fumigatus was isolated and it was not detected in three tissues from which gliotoxin-producing isolates of A. fumigatus were obtained. The ability of this pathogenic fungus to produce this immunomodulating compound in naturally infected turkeys provides further evidence that gliotoxin may be involved in the pathogenesis of the disease, aspergillosis of turkeys. Disclaimer: Names are necessary to report factually on available data; however, the USDA neither guarantees nor warrants the standard of the products, and the use of the name by USDA implies no approval of the product to the exclusion of others that may also be suitable.     

The effects of dsRNA mycoviruses on growth and murine virulence of Aspergillus fumigatus

Some isolates of the opportunistic human pathogenic fungus Aspergillus fumigatus are known to be infected with mycoviruses. The dsRNA genomes of two of these mycoviruses, which include a chrysovirus and a partitivirus, have been completely sequenced and an RT-PCR assay for the viruses has been developed. Through curing virus-infected A. fumigatus isolates by cycloheximide treatment and transfecting virus-free isolates with purified virus, as checked by RT-PCR, isogenic virus-free and virus-infected lines of the fungus were generated whose phenotypes and growth have been directly compared. Mycovirus infection of A. fumigatus with either the chrysovirus or the partitivirus resulted in significant aberrant phenotypic alterations and attenuation of growth of the fungus but had no effect on susceptibility to common antifungals. Chrysovirus infection of A. fumigatus caused no significant alterations to murine pathogenicity.     

Studies on clinical signs and haematological alterations in pneumonic aspergillosis due to Aspergillus flavus in Japanese quail

Intratracheal inoculation of 2-week old quail chicks with Aspergillus flavus spores resulted in the development of clinical signs within 24 h of infection. These were characterized by dullness, depression, anorexia, accelerated breathing, gasping and prostration leading to death. These signs continued up to 7 days followed by considerable decrease in the intensity of the symptoms as well as number of birds showing clinical signs. Mortality occurred primarily in the first week with a majority of the birds dying from 2–4 days after infection. The overall mortality during a 6-week observation period was 25%. The average body weight of the infected chicks was slightly lower than that of controls; the difference being significant at 2, 3 and 42 days post-infection. There was no appreciable difference in the mean values of haemoglobin, packed cell volume and total erythrocyte count between the infected and control chicks at any stage of infection, but total leucocyte count revealed a significant increase (p<0.05) from 3–7 days post-infection. This was due to increase in the percentage of heterophils and decrease in lymphocytes.     

Serum and liver zinc,copper, and iron in chicks infected withEimeria acervulina orEimeria tenella

Two-wk-old broiler chicks were inoculated via crop intubation withEimeria acervulina at two doses: 10⁵ or 10⁶ sporulated oocysts/bird or withEimeria tenella at a dose of 10⁵ sporulated oocysts/bird. Serum and liver samples were collected on days 3 and 6 post-inoculation (PI). There were no significant changes in serum or liver zinc, copper, and iron concentrations in any of the infected groups by 3 d PI. However, on d 6, PI serum protein was significantly reduced in all of the infected groups compared to their pair-fed controls. The chicks infected withE. tennella had significantly reduced serum zinc (1.20 vs 1.77 μg/mL) and iron (0.44 vs 1.28 μg/mL) concentrations and significantly elevated serum copper (0.28 vs 0.17 μg/mL) and ceruloplasmin levels (20.33 vs 11.11 μg/mL) compared to their pair-fed counterparts. Those chicks infected withE. acervulina (10⁶ oocysts/bird) exhibited significantly reduced serum iron concentration by 6 days PI (0.90 vs 1.14 μg/mL). Liver zinc was significantly increased in the chicks infected withE. tenella (349 vs 113 μg/g dry liver wt), as was copper (24 vs 19 μg/g), whereas liver iron concentration was significantly reduced (172 vs 243 μg/g) compared to pair-fed controls. At both dose levels, the chicks infected withE. acervulina exhibited a significant reduction in liver iron by 6 d PI. Hepatic cytosol metals generally reflected whole tissue levels. Metallothionein (MT)-bound zinc was significantly elevated in the chicks infected withE. tenella. Iron bound to a high molecular weight, heat-stable protein fraction (presumably cytoplasmic ferritin) was significantly reduced in chicks infected withE. acervulina, as well as those infected withE. tenella. Collectively, the changes in serum zinc, copper, and iron concentrations, as well as the changes in hepatic zinc and MT-zinc concentrations in the chicks infected withE. tenella were similar to changes evoked during an acute phase response to infection. It is possible that a secondary bacterial infection or inflammation stemming from erosion of the lining of the cecum may play a role in the response of trace element metabolism to theE. tenella infection. Mentions of a trademarkr, proprietary product or specific equipment does not consitute a guarantee or warranty by the US Department of Agriculture and does not imply its approval to the exclusion of other products.     

Airborne Aspergillus fumigatus conidia: a risk factor for aspergillosis

Aspergillus fumigatus is an opportunistic fungal pathogen that causes invasive aspergillosis, a usually fatal infection. The disease has risen in prominence in recent years due to the increasing numbers of severely immunocompromised patients becoming infected. The fungus is ubiquitous in the environment, producing large numbers of conidia that are dispersed in the air. Humans inhale numerous conidia everyday, but infections are not seen in healthy individuals. As inhalation of conidia is the main route of infection, considerable efforts are required to prevent infection in susceptible patients. This review summarises the current knowledge on airborne concentrations of A. fumigatus conidia, their background levels in outdoor air and seasonal distribution patterns. New and established methods of air sampling for airborne A. fumigatus conidia are discussed. Common environmental sources of the fungus are reviewed, including its presence in compost heaps. Finally, the lack of stringent guidelines on the monitoring and control of airborne A. fumigatus concentrations in hospitals is discussed.