Occurrence and species distribution of pathogenic Mucorales in unselected soil samples from France.

Mucormycosis is a life-threatening invasive fungal disease that affects a variety of patient groups. Although Mucorales are mostly opportunistic pathogens originating from soil or decaying vegetation, there are currently few data on prevalence of this group of fungi in the environment. The aim of the present study was to assess the prevalence and diversity of species of Mucorales from soil samples collected in France. Two grams of soil were homogenized in sterile saline and plated on Sabouraud dextrose agar and RPMI agar supplemented with itraconazole or voriconazole. Both media contained chloramphenicol and gentamicin. The plates were incubated at 35 ± 2 °C and checked daily for fungal growth for a maximum of 7 d. Mucorales were subcultured for purity. Each isolate was identified phenotypically and molecular identification was performed by ITS sequencing. A total of 170 soil samples were analyzed. Forty-one isolates of Mucorales were retrieved from 38 culture-positive samples. Among the recovered isolates, 27 Rhizopus arrhizus, 11 Mucor circinelloides, one Lichtheimia corymbifera, one Rhizopus microsporus and one Cunninghamella bertholletiae were found. Positive soil samples came from cultivated fields but also from other types of soil such as flower beds. Mucorales were retrieved from samples obtained in different geographical regions of France. Voriconazole-containing medium improved the recovery of Mucorales compared with other media. The present study showed that pathogenic Mucorales are frequently recovered from soil samples in France. Species diversity should be further analyzed on a larger number of soil samples from different geographic areas in France and in other countries.

Molecular monitoring of fungal communities in air samples by denaturing high-performance liquid chromatography (D-HPLC).

AIMS: To describe a new molecular technique for the assessment of fungal diversity in the air. METHODS AND RESULTS: Air samples were collected every week in a henhouse in France during a 15-week period. After air sampling, the collecting membrane was diluted, and the liquid was used for subsequent cultivation and molecular analysis: PCR-temperature temporal gradient electrophoresis (TTGE), which has already been used for the identification of fungal species in air samples and PCR-denaturing high-performance liquid chromatography (D-HPLC), a new technique for the analysis of complex microbial populations. D-HPLC profiles were reproducible from run-to-run, and several fungal organisms could be identified at the species level by sequencing. CONCLUSIONS: PCR-D-HPLC enabled the identification of fungal species (both Ascomycota and Basidiomycota) that may be encountered in air. The new technique allowed the detection of more fungal species than did the PCR-TTGE technique. However, some fungal species were detected only by PCR-TTGE, suggesting that PCR-D-HPLC and PCR-TTGE are complementary. SIGNIFICANCE AND IMPACT OF THE STUDY: PCR-D-HPLC represents a considerable saving in time over currently available procedures for detection and identification of fungal organisms in air. However, the fungal diversity detected by PCR-D-HPLC or by PCR-TTGE was lower than that revealed by culture.

Effects of conidia of various Aspergillus species on apoptosis of human pneumocytes and bronchial epithelial cells.

Aspergillus species can cause mycoses in human and animals. Previously, we demonstrated that A. fumigatus conidia from a human isolate inhibited apoptosis in human pneumocytes and bronchial epithelial cells. In the current study, we studied the effects of A. fumigatus conidia non-human origin and A. flavus, A. nidulans, A. niger and A. oryzae conidia on human cells apoptosis. Human pneumocytes or bronchial epithelial cells were simultaneously exposed to apoptotic inductors and aspergilli conidia. The cell cultures were analyzed by flow cytometry, immunoblotting, and examination of nuclear morphology. Similar to A. fumigatus conidia, A. flavus conidia inhibited cellular apoptosis while A. nidulans, A. niger and A. oryzae conidia did not affect apoptosis. We further studied the species specificity of conidia: there were no differences in the inhibition of apoptosis by A. fumigatus conidia from either human or bird isolates. In order to determine whether the inhibition of apoptosis by conidia is limited to certain strains, the effect on human cell apoptosis of different A. fumigatus human clinical isolates and A. fumigatus of environmental origin was evaluated. All A. fumigatus isolates inhibited apoptosis; an anti-apoptotic factor was released by conidia. For TNF-induced apoptosis, the anti-apoptotic effect of conidia of all isolates was found to be associated with a reduction of caspase-3 in human cells. The results suggest that suppression of apoptosis may play a role in reducing the efficacy of host defense mechanisms during infection with Aspergillus species.

Evolution of the environmental contamination by thermophilic fungi in a turkey confinement house in France.

Fungal species constitute a major part of environmental contaminants in facilities where animals are housed. The present investigation was aimed at describing the relative abundances of fungal species and their concentrations in a turkey confinement house in France. Fungal cultures from poultry feed, litter, and air were undertaken every week throughout the 16-wk period of breeding. The incubation temperature of 40 degrees C was selected to isolate thermophilic fungal species (especially Aspergillus spp. and Candida albicans) that are potentially pathogenic for birds. The 2 species Aspergillus fumigatus and Aspergillus flavus were recovered at a mean of 10.5 and 37.0 cfu/m(3) of air sampled, respectively. Individual samplings yielded concentrations of up to 150.0 cfu/m(3) for A. flavus in the first weeks of the investigation. Other fungal species were recovered at a mean of 18.9 cfu/m(3) (maximum 36.3 cfu/m(3)) in the air. The yeast C. albicans was first detected at wk 4 from litter samples and at wk 7 from poultry feed. Densities of C. albicans remained very high in litter samples (63.2 cfu/g) even after new litter was added at wk 10. To analyze the genetic polymorphism of A. fumigatus, the most pathogenic mold in birds, a total number of 198 isolates (134 from air, 34 from litter, and 30 from feed samples) were genotyped using 2 polymorphic microsatellite markers. More than half (42 out of 73, 57.5%) of the genotypes were detected only once. This finding suggests that the contamination of the breeding environment is not due to a single source and confirms the very high genetic diversity of environmental A. fumigatus isolates. As during the study period, no outbreak of fungal infections occurred; the levels of fungal contaminations reported here do not seem sufficient, at least alone, to trigger fungal infections.

Prevalence of Cryptosporidium infection in calves in France.

Two multicentre surveys were conducted in France to estimate the prevalence of Cryptosporidium infection in calves using qualitative ELISA for detection of Cryptosporidium coproantigens and oocysts. The first survey involved 4-12-day-old calves in six dairy-calf distribution centres, collecting calves from seven Administrative Regions (Aquitaine, Bretagne, Franche-Comté, Lorraine, Normandie, Nord, Pays de Loire). For each region, 20 calves were selected every month for 12 consecutive months (October 1995-September 1996). Prevalence of Cryptosporidium infection was 17.9% (Confidence Intervals (C.I.) 95%=[16.1%; 19.8%]) among the 1628 selected calves, of which only 5.3% had diarrhoea. The second survey conducted between November 1995 and May 1996 involved 4-21-day-old calves examined by veterinary practitioners who selected 189 livestock farms of dairy- or suckler-type in ten Administrative Departments (Allier, Cantal, Creuse, Doubs, Ille-et-Vilaine, Maine-et-Loire, Manche, Pas-de-Calais, Saône-et-Loire, Vendée). Cryptosporidia were detected in 105 (55.6%) of the farms. Among the 440 calves examined, of which 398 (90.5%) presented diarrhoea, cryptosporidia were found in 191 animals, i.e. a prevalence of 43.4% (C.I. 95%=[38. 8%; 48.0%]). Breed of calves and type of housing had very little impact on prevalence in this survey. Some regional variations could be noticed, even if cryptosporidia infection is widespread. Monthly variations could be related to seasonal peaks in calving with a lower infection rate during summer.

Role of Cryptosporidium parvum as a pathogen in neonatal diarrhoea complex in suckling and dairy calves in France.

This study was carried out to find the importance of Cryptosporidium parvum in diarrhoea of neonatal calves in two types of breeding - suckling and dairy calves - in France. Different agents causing neonatal diarrhoea, E. coli, rotavirus, coronavirus, Salmonella and Cryptosporidium were systematically researched in faeces. 1. Suckling calves: In 40 livestock farms selected for diarrhoea, 311 calves 4 to 10 days old which had diarrhoea for less than 24h or no diarrhoea, were included in the study. A prophylaxis of neonatal diarrhoea had been carried out in 21 of the 40 livestock farms. On D0 (inclusion day), the mean age was 6 days, 82% presented a good initial general condition and 76.2% had a good appetite; 48.6% were diarrhoeic but 91.3% presented no sign of dehydration. Only 6.1% were infected by E. coli K99, 14.3% by rotavirus, 6.8% by coronavirus, 0.3% by Salmonella but 50% excreted C. parvum oocysts. This later percentage increases up to 84% and 86% by D3 and D7, respectively . We note that 16% of the 4-day-old calves on D0 are excreting oocysts and this percentage increases as a function of the age of the calf on D0 to reach 90% to 95% by the age of 8 days. 10 out of 12 dead calves excreted C. parvum oocysts. From D0 to D14 the other pathogen agents show a relative or a decreasing stability. 2. Dairy calves: 382 calves which had diarrhoea for less than 24 h or no diarrhoea, aged 8 to 15 days coming from six industrial livestock farms were included in the study. On D0, 99% of the calves presented a good initial general condition, 99.7% had a good appetite and no calf was dehydrated. At this date (D0), 16.8% of the calves excreted cryptosporidia. This percentage increases up to 23% and 51.8% on D3 and D8, respectively, then decreases to 31.9% on D14. The pressure of the other pathogenicagents remains relatively stable, excepted for rotavirus on D7 (from 9.9% on D0 to 27.2% on D7, then 12.6% on D14) which does not explain the concomitantpeak in diarrhoea because the infection by rotavirus on D7 is more frequent in non-diarrhoeic calves than in diarrhoeic calves. Our results show that Cryptosporidium prevalence is higher in suckling than in dairy calves and C. parvum constitutes actually in both cases the major aetiological agent of neonatal diarrhoea.

Epidemiological analysis of Malassezia pachydermatis isolates by partial sequencing of the large subunit ribosomal RNA.

The opportunistic yeast Malassezia pachydermatis is commonly recovered from both normal and diseased skin of warm-blooded animals. The diversity of M pachydermatis isolates obtained from a wide range of hosts was investigated by the partial sequencing of the large subunit (LSU) ribosomal RNA. Among 100 isolates examined, seven types (Ia-Ig) were discriminated on the basis of nucleotide sequence diversity. The seven types differed by one to five mutations, all of them corresponding to transitions. The predominant sequence, type Ia, appeared to be ubiquitous since it was observed in isolates recovered from domestic and wild carnivora, from a monkey and from man. In contrast the sequence types Ic, Id and Ig seemed to be more host-specific; they included isolates recovered exclusively from rhinoceros, dogs and ferrets, respectively. None of the seven sequence types correlated with isolation from healthy skin or a particular lesion (otitis externa or other dermatitis). The study indicated that the skin of an animal may be colonised by more than one type of M pachydermatis.

[The role of companion animals in the spread of zoonoses of parasitic origin]. / Rôle des animaux de compagnie dans la dispersion des zoonoses d'origine parasitaire.

This paper provides parasitological data necessary to the epidemiology of the major parasitic zoonoses transmitted by companion animals. Possible modes of contamination of humans are reviewed, along with various factors linked to parasites, host animals and human beings in the spread of zoonoses.

Fungal flora on cutaneous and mucosal surfaces of cats infected with feline immunodeficiency virus or feline leukemia virus.

OBJECTIVE: To compare cutaneous and mucosal mycoflora in cats infected with FIV or FeLV with that in noninfected cats. ANIMALS: 85 client-owned cats; 24 seropositive for FIV, 10 seropositive for FeLV, 1 seropositive for both viruses, and 50 seronegative for both viruses. PROCEDURE: Cutaneous specimens were obtained from the coat and external acoustic meatus (ear canal) and mucosal specimens from the oropharynx and rectum. Fungi were isolated from specimens, using Sabouraud dextrose agar incubated at 27 or 37 C for cutaneous and mucosal specimens, respectively. RESULTS: Fungal colonies were cultured from at least 1 specimen from 83 of 85 (97.6%) cats. The most common fungal isolates were Aspergillus spp (cultured from 59.3% of all specimens), Penicillium spp (50.0%), Cladosporium spp (44.2%), Scopulariopsis spp (41.8%), and lipophilic yeasts of the genus Malassezia (31.4%). A greater diversity of fungal genera was isolated from retrovirus-infected cats, and Malassezia spp were more commonly recovered from these cats, compared with noninfected cats. Candida albicans, Cryptococcus neoformans, and dermatophytes (eg, Microsporum canis) were rarely isolated from any cat. Significant differences in frequency of isolation of C. neoformans and dermatophytes were not found between infected and noninfected cats. CONCLUSIONS AND CLINICAL RELEVANCE: Cats infected with FIV or FeLV may have a greater diversity of cutaneous and mucosal mycoflora than noninfected cats. However, infected cats may be no more likely than noninfected cats to expose humans to zoonotic fungi such as C. albicans, C. neoformans, and M. canis.

Subcutaneous Taenia crassiceps cysticercosis in a dog.

Taenia crassiceps larvae, typically found in rodents, were recovered from fluctuant subcutaneous cystic lesions in a dog. Differential diagnosis included Taenia crassiceps cysticercosis, other metacestode infections, and pseudotumoral skin diseases.

Efficacy of halofuginone lactate in the prevention of cryptosporidiosis in suckling calves.

The efficacy of halofuginone lactate in the prevention of cryptosporidiosis in suckling calves was evaluated in a multicentre, control versus placebo, randomised, double-blind clinical trial. Seventy-eight six- to 48-hour-old calves were treated daily with 120 microg/kg bodyweight of halofuginone lactate administered orally for seven consecutive days, while 80 calves received a placebo. Faecal samples were collected on the first day of dosing and four, seven, 14 and 21 days later, and Cryptosporidium parvum oocysts were counted and faecal indices for diarrhoea were determined after a clinical examination. An analysis of variance for repeated measurements showed a highly significant difference in favour of halofuginone lactate for both the oocyst counts (P=0.0002) and the faecal diarrhoea indices (P=0.0001) throughout the trial. The difference was greatest after seven days, when the mean oocyst count of the placebo group was 2.5 times and its mean faecal index was twice the mean of the halofuginone lactate group. One day after the end of the treatment the calves which received halofuginone lactate excreted 44 per cent fewer C parvum oocysts and 44 per cent fewer of them had diarrhoea. The reduction was even greater (65 per cent) when liquid diarrhoea was assessed, with 32.5 per cent of the calves in the placebo group having liquid diarrhoea compared with 11.5 per cent in the halofuginone lactate group. The treatment was well tolerated and easily administered.

Evaluation of the efficacy of oral lufenuron combined with topical enilconazole for the management of dermatophytosis in catteries.

The efficacy of oral lufenuron, a chitin synthetase inhibitor, combined with topical enilconazole, was evaluated for the management of Microsporum canis infection in 100 cats housed in two catteries in France. The cats were treated with weekly rinses with enilconazole (0.2 per cent) for four weeks and, in each cattery, one group (A) was also treated with micronised griseofulvin (25 mg/kg administered orally twice a day for five weeks) and a second group (B) was treated with 60 mg/kg lufenuron administered orally once on day 0 and again after 30 days. All the cats were examined individually for cutaneous lesions and mycological cultures were made when the treatment began and after 15, 30, 60 and 90 days. In the first cattery, the cats' clinical scores after 30 and 60 days were significantly lower in group B than in group A. In both catteries and both treatment groups, the mean number of fungal colonies decreased rapidly during the first 15 days of treatment, remained stable for the following 45 days but increased from day 60 to the end of the experiment on day 90.

[Invasive cysticercosis due to Taenia crassiceps in an AIDS patient]. / Cysticercose envahissante à Taenia crassiceps chez un patient atteint de sida.

A first case is reported from France of a subcutaneous and muscular tissues invasion by proliferative larval forms of Taenia crassiceps in an AIDS-patient with severe immunodeficiency. These larvae usually develop in rodents whereas the adult form of the cestode is commonly found in foxes. Authors emphasize the differential diagnosis from other human subcutaneous metacestodoses such as T. solium cysticercosis, cenurosis and hydatidosis. The proliferative aspect of the larvae, the recurrence of lesions after surgical removal and the possible invasion of other tissues are elements of a poor prognosis.