Nocardia asteroides as a cause of pneumonia in a buffalo calf

Pulmonary nocardiosis has been described in a four-month-old male buffalo calf (Bubalus bubalis) with a history of respiratory distress lasting ten days. Microscopic examination of tissue imprints from the infected lungs revealed numerous Gram-positive, acid-fast, fine, branched, filamentous organisms which were morphologically identical with those of Nocardia spp. Cultures of pneumonic lung specimens on plain Sabouraud medium yielded pure growth of Nocardia asteroides at 37 degrees C. Lungs from ten healthy buffalo gave negative results for bacteria, suggesting that N. asteroides does not occur as a commensal in the pulmonary tissues. Epidemiological investigation established the source of infection in the immediate environment of the animal. It is emphasised that N. asteroides may be considered to be a respiratory pathogen and should be included in differential diagnosis of pulmonary diseases.     

Molecular study of persistence of Nocardia asteroides and Nocardia otitidiscaviarum strains in patients with long-term nocardiosis

Three Nocardia asteroides and five Nocardia otitidiscaviarum isolates recovered from three patients with long-term nocardiosis were compared by random amplification of polymorphic DNA fingerprinting, antibiotic susceptibility testing, and enzymatic characterization. Results obtained with primer DKU49 (P. Palittapongarnpim, S. Chomyc, A. Fanning, and D. Kunimoto, J. Infect. Dis. 167:975-978, 1993) provide evidence that patient A was infected by two N. asteroides strains during a single episode of nocardiosis and that patients B and C remained infected by the same strain, respectively. Resistance to minocycline that was present in the first isolate recovered from patient B reverted to intermediate resistance in the second isolate and reverted to susceptibility in the third isolate. Resistance to penicillin G and beta-lactams was acquired by the second isolate obtained from patient C.     

Ribotyping: a tool for taxonomy and identification of the Nocardia asteroides complex species

Ribotyping has been evaluated as a taxonomic tool for studying bacteria belonging to the Nocardia asteroides complex. The probe used was obtained by PCR from DNA extracted from the type strain Nocardia asteroides ATCC 19247, a sequence that codes partly for 16S rRNA. Interpretation of hybridization after EcoRI restriction of total DNA of 21 strains of the N. asteroides complex allowed for the identification of only four different ribotypes: two related to N. asteroides sensu stricto, one related to N. farcinica and one related to N. nova. These results confirm the classification established by Tsukamura in 1982 (M. Tsukamura, Microbiol. Immunol. 26:1101-1119, 1982) as well as the heterogeneity of N. asteroides sensu stricto species. Ribotyping may also provide a major identification technique for the species belonging to the N. asteroides complex.     

Late lateralizing and localizing EEG features of scalp-recorded temporal lobe seizures

The fungicidal properties of the synthetic peptide D4E1 were studied with nongerminated and germinating conidia of Aspergillus flavus, Aspergillus fumigatus, Aspergillus niger, Fusarium moniliforme, and Fusarium oxysporum. The minimal lethal concentrations (MLC) needed to kill 100% of germinating conidia of A. fumigatus, A. flavus, and A. niger were 12.5, 12.5, and 25 microM, respectively. The MLC value for nongerminated and germinating conidia of both Fusarium spp. was 3.0 microM. Except for A. fumigatus, D4E1 was inactive against the nongerminated conidia of the Aspergillus spp. Physicochemical studies showed D4E1 complexed with ergosterol, a sterol present in conidial walls. Cholesterol, present in nongerminated conidia of F. moniliforme, had a greater affinity for D4E1 than did ergosterol. D4E1 was more resistant to fungal and plant protease degradation than the natural peptide, cecropin A. These in vitro results suggest D4E1 is a candidate for transgenic expression in plants to enhance host resistance to fungal infection.     

Opacification of Middlebrook agar as an aid in distinguishing Nocardia farcinica within the Nocardia asteroides complex

Among 58 aerobic actinomycetes isolated from different sources and geographical locations, none of 23 Nocardia asteroides isolates, at 18 N. farcinica isolates, 1 of 5 N. otitidiscaviarum isolates, and 1 of 4 Rhodococcus species isolates opacified Middlebrook 7H10 medium. Within the N. asteroides complex, this characteristic, together with growth at 45 degrees C and resistance to each of erythromycin, cefotaxime, and tobramycin, provides a simple means of distinguishing N. farcinica from N. asteroides.     

Oxygen requirements of Aspergillus species

The growth of 24 Aspergillus isolates at low oxygen tensions was assessed. Isolates selected included A. fumigatus (10), A. terreus (6), A. niger (6), A. nidulans (1) and A. flavus (1). Three different agar media were used--potato dextrose agar (PDA), pH 5.6; brain heart infusion (BHI), pH 7.4; and a specially developed medium (Hall's) containing resazurin--with oxygen concentrations of 0, 0.025, 0.1, 0.5 and 2.5%. The CO2 concentration was 5%. Agar plates were inoculated with 2 x 10(7) conidia/ml, loaded into jars and flushed with a special gas mixture at 37 degrees C. The plates were inspected at intervals of 3, 5 and 10 days. On Hall's medium, none of the isolates grew at oxygen concentrations of 0 or 0.025%, but 21 (88%) of 24 grew at 0.1%. On PDA and BH1, all 14 isolates tested grew at oxygen concentrations of 0.5 and 2.5%. Three of these 14 conidiated on PDA at oxygen 0.5% and 12 of 14 conidiated on PDA at oxygen 2.5%. None grew without oxygen on these media. Thus, pathogenic Aspergillus spp. are capable of growth at low oxygen tensions, and this may have implications for pathogenicity and antifungal activity.     

Sordarins: in vitro activities of new antifungal derivatives against pathogenic yeasts, Pneumocystis carinii, and filamentous fungi

GM 193663, GM 211676, GM 222712, and GM 237354 are new semisynthetic derivatives of the sordarin class. The in vitro antifungal activities of GM 193663, GM 211676, GM 222712, and GM 237354 against 111 clinical yeast isolates of Candida albicans, Candida kefyr, Candida glabrata, Candida parapsilosis, Candida krusei, and Cryptococcus neoformans were compared. The in vitro activities of some of these compounds against Pneumocystis carinii, 20 isolates each of Aspergillus fumigatus and Aspergillus flavus, and 30 isolates of emerging less-common mold pathogens and dermatophytes were also compared. The MICs of GM 193663, GM 211676, GM 222712, and GM 237354 at which 90% of the isolates were inhibited (MIC90s) were 0.03, 0.03, 0.004, and 0.015 microg/ml, respectively, for C. albicans, including strains with decreased susceptibility to fluconazole; 0.5, 0.5, 0.06, and 0.12 microg/ml, respectively, for C. tropicalis; and 0.004, 0.015, 0.008, and 0.03 microg/ml, respectively, for C. kefyr. GM 222712 and GM 237354 were the most active compounds against C. glabrata, C. parapsilosis, and Cryptococcus neoformans. Against C. glabrata and C. parapsilosis, the MIC90s of GM 222712 and GM 237354 were 0.5 and 4 microg/ml and 1 and 16 microg/ml, respectively. The MIC90s of GM 222712 and GM 237354 against Cryptococcus neoformans were 0.5 and 0.25 microg/ml, respectively. GM 193663, GM 211676, GM 222712, and GM 237354 were extremely active against P. carinii. The efficacies of sordarin derivatives against this organism were determined by measuring the inhibition of the uptake and incorporation of radiolabelled methionine into newly synthesized proteins. All compounds tested showed 50% inhibitory concentrations of <0.008 microg/ml. Against A. flavus and A. fumigatus, the MIC90s of GM 222712 and GM 237354 were 1 and 32 microg/ml and 32 and >64 microg/ml, respectively. In addition, GM 237354 was tested against the most important emerging fungal pathogens which affect immunocompromised patients. Cladosporium carrioni, Pseudallescheria boydii, and the yeast-like fungi Blastoschizomyces capitatus and Geotrichum clavatum were the most susceptible of the fungi to GM 237354, with MICs ranging from /=2 microg/ml. In summary, we concluded that some sordarin derivatives, such as GM 222712 and GM 237354, showed excellent in vitro activities against a wide range of pathogenic fungi, including Candida spp., Cryptococcus neoformans, P. carinii, and some filamentous fungi and emerging invasive fungal pathogens.     

Expression pattern of aspartic proteinase antigens in aspergilli

Sporulating colonies of Aspergillus fumigatus, A. flavus, and A. niger were subjected to immunofluorescence using specific polyclonal antibodies against the aspergillopepsin PEP (EC 3.4.23.18), a secretory aspartic proteinase produced by A. fumigatus. The proteinase antigen was found mainly in developing conidiophores of aspergilli, in submerged mycelia and on the tips of growing aerial mycelia. Mature aerial hyphae and spores showed no immunofluorescence at all. Sporulating conidiophores revealed only weak activity in A. fumigatus and A. flavus. The distinct pattern of expression of the aspartic proteinase antigens suggests a role for such enzymes in the growth of hyphae and the development of conidiophores and thus for the sporulation process in aspergilli.     

Corneal ulcer caused by Nocardia asteroides in a patient with leprosy

Nocardia asteroides is a rare cause of keratitis usually associated with trauma. We report a case of corneal ulceration caused by N. asteroides in a patient with leprosy. This is the first case report of nocardial keratitis from Southeast Asia. The diminished corneal sensation in a patient with leprosy could be a predisposing factor for development or exacerbation of corneal ulceration.     

Influence of zinc oxide on Aspergillus species: a possible cause of local, non-invasive aspergillosis of the maxillary sinus

Recently the appearance of radiopaque 'concrements' in the maxillary sinus was reported. These radiodense objects could be identified as root-filling material for teeth of the upper jaw containing zinc oxide. This suggested that excess root-filling material containing zinc oxide in the maxillary sinus could favour the formation of a local, non-invasive aspergillosis. To verify this hypothesis in vitro, we tested the influence of zinc oxide on Aspergillus fumigatus, A. flavus, A. terreus, A. nidulans, A. niger and A. niveus. A geometric series of zinc oxide diluted in 0.1 N H2SO4 was used. Czapek-Dox agar was inoculated with the six Aspergillus species. Circular cavities stamped into the centre of each agar plate were filled with an equal amount of the dilutions and all plates were then incubated for 7 days at 37 degrees C. Readings were taken on days 3, 5 and 7. Soluble zinc oxide promoted the growth of all the tested Aspergillus species, the effect diminishing with decreasing concentration. This effect could be observed on all days of measurement. Only with A. niger was stimulation of growth minimal, probably because this species already exhibits a high growth rate on Czapek-Dox agar. Therefore, overfilling of maxillary teeth with a zinc oxide-containing root-filling material may favour the formation of local aspergillosis.     

Cardiac tamponade due to Nocardia asteroides

Pericarditis with hemodynamic compromise is a rare manifestation of infection with Nocardia asteroides. To our knowledge, only six cases have been reported previously. In contrast to other cases of pericardial disease due to Nocardia, culture of the pericardial fluid in our case was negative while culture of pericardial tissue led to the diagnosis. Surgical intervention and appropriate antibiotic therapy are essential in the treatment of Nocardia pericarditis.     

Brasilinolide A, a new macrolide antibiotic produced by Nocardia brasiliensis: producing strain, isolation and biological activity

A new 32-membered macrolide antibiotic, brasilinolide A was isolated from the fermentation broth of Nocardia sp. IFM 0406. The producer was identified as Nocardia brasiliensis. The antibiotic was only active against Aspergillus niger, but not active against other fungi including yeasts as well as other filamentous like fungi and bacteria. Brasilinolide A exerted an immunosuppressive activity in the assay system of a mixed lymphocyte reaction (MLR).     

Allergic fungal sinusitis: clinicopathological characteristics

Allergic fungal sinusitis is a comparatively new disease entity in paranasal sinus mycoses. It is not a very rare condition, but diagnosis is difficult to establish. Of 28 consecutive cases of allergic nasal polyposis during a 2-year period, 11 patients had allergic fungal sinusitis and the diagnosis was based on the presence of type I hypersensitivity, eosinophilic mucus without tissue invasion of fungi on histopathology and detection of septate hyphae on direct microscopy. On culture, Aspergillus flavus was isolated from nine patients and A. fumigatus and A. niger from one patient each. Among patients with allergic fungal sinusitis, five had asthma, four had proptosis, of whom two had impaired vision, and all 11 patients had nasal obstruction. Eight patients described a history of recurrence. All patients underwent surgical clearance of the diseased sinuses and were given post-operative topical steroids to prevent recurrence.     

Funding nursing education: the Medicare dilemma. Interview by Cheryl Slagle King

A total of 110 patients of symptomatic otomycosis was investigated, prospectively. Aural swabs were collected on 1st, 7th and 14th day and examined by direct microscopy and culture for fungi. Of these, 80 patients found to be having pure fungal infection, were taken up for mycological and therapeutic study. Fungi belonging to genus Aspergillus were isolated in 76 (95.0%) patients of which Aspergillus niger was the commonest isolate in 46 (57.5%), followed by A. flavus in 27 (33.7%), A. fumigatus in 3 (3.7%), Candida species in 3 (3.7%) and Mucor in 1 (1.2%). The patients were of all age groups but majority were between 21 and 30 years and the male-female ratio was equal. Of the total of 40 male patients, twenty-one were Sikhs using turban. Before developing the symptoms, forty five patients used oil, mixture of oil and garlic juice, antibiotics, steroids, antiseptics or wax solvent as ear drops. Only two patients were diabetic. No patient had fungal infection elsewhere in the body. The patients were called for regular follow-up for three weeks. In forty cases mercurochrome was applied as the antifungal agent after cleaning the external auditory canal, in twenty-three clotrimazole and in rest of the seventeen patients miconazole was used. On 7th day, only 11 (13.7%) patients grew different fungi in culture. They became symptom-free on 14th day and no fungal material could be seen on otoscopy, direct microscopy or culture. Mercurochrome was found to be most effective in these patients.     

Disseminated infection due to an unusual strain of Nocardia farcinica

Nocardia farcinica, isolated by Nocard in 1888, has recently been redefined as a separate species to asteroides and has been recognised as human pathogen since 1975. Following airborne contamination, lung infection is most usual with frequent dissemination especially to central nervous system. Usually drug resistance makes treatment difficult. The authors describe the first case of Nocardia farcinica disseminated infection in France with lung primarily affection, empyema and encephalitis.     

Genetic diversity among clinical and environmental isolates of Aspergillus fumigatus

To determine if cases of invasive aspergillosis (IA) were caused by strains of Aspergillus fumigatus with unique characteristics, strains from immunosuppressed patients with IA were compared to strains obtained from sputa of patients with cystic fibrosis and to strains from the environment. An extremely high genomic diversity was observed among the 879 strains typed by Southern blotting with a retrotransposon-like element from A. fumigatus (C. Neuvéglise, J. Sarfati, J. P. Latgé, and S. Paris, Nucleic Acids Res. 24:1428-1434, 1996). Analysis of Southern blot hybridization patterns showed the absence of clustering between environmental isolates and clinical isolates from patients with IA or cystic fibrosis. In addition, strains could not be clustered depending on their geographical location. This study implies that practically any strain of A. fumigatus is potentially pathogenic and can provoke a case of IA when it encounters a favorable environment in an immunosuppressed host.     

Determination of alprazolam and alpha-hydroxyalprazolam in human plasma by gas chromatography/negative-ion chemical ionization mass spectrometry

A sensitive and specific method was developed for the determination of alprazolam and its major metabolite alpha-hydroxyalprazolam in plasma. After the addition of deuterium-labeled internal standards, plasma samples were buffered to pH 9 with 1 ml of saturated sodium borate buffer, extracted with toluene-methylene chloride (7:3) and evaporated to dryness. The residues were treated with N,O-bis(trimethylsilyl)trifluoroacetamide containing 1% of trimethylchlorosilane and analyzed on a Finnigan-MAT mass spectrometer operated in the negative-ion chemical ionization mode with methane as the reagent gas. Chromatographic separation was achieved on a Restek-200 capillary column using hydrogen as the carrier gas. The assay was linear from 0.25 to 50 ng ml-1 for both compounds. The intra-assay precision for alprazolam was 16.1% at 0.5 ng ml-1 and 4.6% at 50 ng ml-1 and that for alpha-hydroxyalprazolam was 15.8% at 0.5 ng ml-1 and 4.2% at 50 ng ml-1. The method was used to determine alprazolam and alpha-hydroxyalprazolam in human plasma samples collected after a single 2 mg oral does of alprazolam. A peak concentration of 32.9 ng ml-1 of alprazolam was detected at 1 h following the dose.     

Reduction in the growth of non-dermatophyte moulds by itraconazole: evaluation by corneofungimetry assay

The activity of itraconazole against fungi of dermatological importance was studied using corneofungimetry performed on four test organisms: Aspergillus flavus, Fusarium oxysporum, Scopulariopsis brevicaulis and Scytalidium dimidiatum. The human stratum corneum used in the bioassay was harvested from healthy volunteers and from patients undergoing treatment with itraconazole 200 mg daily for at least 2 weeks. A fungal mycelium grew on all untreated stratum corneum samples within 1 week. The pattern and extent of growth varied greatly among the test organisms. An inhibitory effect was observed on the itraconazole-treated samples. However, the level of inhibition was higher than that expected by calculating itraconazole minimum inhibitory concentrations in a reference susceptibility test. It is concluded that some non-dermatophyte moulds of medical importance can grow by forming hyphae on human stratum corneum. However, the process is partly inhibited by some unknown mechanisms originating from the stratum corneum itself. It is further inhibited by the oral administration of itraconazole. This global effect might be the result of the complementation inhibition due to the drug and/or some unknown components of the stratum corneum.     

Deoxyribonucleic acid reassociation in the classification of the 'rhodochrous' complex and allied taxa

The degree of binding was determined between DNA preparations from gordonae and rhodochrous strains and uracil-labelled DNA from five reference strains, Nocardia asteroides NK20, N. pellegrino PII, Gordona bronchialis TI, G. terrae T5 and rhodochrous strain R90. Most of the rhodochrous and pellegrino strains fell into one of two genetically homogeneous taxa. The nucleotide sequence homology data also suggested that G. bronchialis, G. rubra, and more equivocally G. terrae, formed distinct species. However, the values for DNA relatedness between these species, and between then and the rhodochrous homology groups, were comparatively low. Only a small degree of nucleotide sequence homology was found between the N. asteroides reference system and the rest of the taxa studied. The nucleotide composition of the DNA preparations from 29 of the 30 test strains fell between 63 and 69 mol% guanine plus cytosine.     

Cloning and sequence analysis of the gene (eprA1) encoding an extracellular protease from Aeromonas hydrophila

A gene (eprA1) encoding the extracellular protease of Aeromonas hydrophila AH1 has been cloned and sequenced. Nucleotide sequence analysis of eprA1 predicted a single open reading frame (ORF) of 1038 bp encoding a 346 amino acid (aa) polypeptide, with a potential 21-aa signal peptide. When the eprA1 gene was expressed in minicells, one major band of approx. 37 kDa was identified, while protease activity staining experiments identified a caseinolytic band of approx. 29 kDa determined by SDS-PAGE analysis of the minicells. The deduced C-terminal aa region (Arg-290 to Gly-313) showed sequence homology to partial C-terminal sequences of other zinc metalloproteases including Penicillium citrinum metalloprotease (PlnC), Aspergillus oryzae metalloprotease (NpII), Aspergillus flavus metalloprotease (MepA), and Aspergillus fumigatus metalloprotease (Mep20), particularly with respect to zinc-binding residues.