Observation et étude expérimentale de mycobactéries atypiques en aquariums d'eau douce et d'eau de mer Observation and experimentatal study of typical mycobacteria in fresh water and salt water aquaria

Résumé

L'eau des aquariums est source de Mycobactéries atypiques qui peuvent être pathogènes pour l'homme et les poissons.

Une étude a été réalisée à l'aquarium du Musée de Zoologie de Nancy. La recherche de Mycobactéries a été effectuée dans 40 aquariums équipés de lampes germicides à UV : 11 bassins étaient alimentés en eau douce et 29 en eau de mer. Deux aquariums non équipés de système de désinfection ont également été analysés, les propriétaires de ces derniers ayant présenté un granulome cutané à M. marinum. L'action des UV sur M. marinum en suspension dans l'eau a été testée expérimentalement dans des bassins d'eau douce peuplés de Cichlidés.

Pour chaque aquarium, un échantillon de 250 ml a été prélevé. Les cultures après décontamination au lauryl sulfate de soude ont été réalisées sur milieu de Loewenstein.

Les résultats indiquent que, quel que soit l'aquarium, la présence de mycobactéries est constante. L'isolement des mycobactéries peut être gêné par la présence d'une flore bactérienne ou fongique importante (≥ 10³ U.F.C./ml).

Les espèces les plus fréquemment isolées sont M. gordonae et M. fortuitum ; M. kansasii et M. marinum ont rarement été isolées (6140 aquariums). Différents facteurs peuvent intervenir sur la sélection des espèces. La salinité de l'eau limite le développement de certaines espèces, alors qu'elle permet la croissance de M. fortuitum. Dans notre étude, la température de l'eau n'a pas été un facteur sélectif. L'utilisation de lampes UV limite le nombre de Mycobactéries. Dans les bassins expérimentaux, les radiations UV se sont révélées très actives sur M. marinum en présence ou en l'absence de poissons. A l'inverse de M. gordonae, M. fortuitum est rarement isolée en présence d'UV. Un nombre important de poissons par aquarium augmente la flore bactérienne et mycobactérienne.

La prévention des infections à Mycobactéries atypiques chez l'homme comme chez les poissons devrait pouvoir être assurée par des mesures d'hygiène élémentaire.

Mots-clés:

• mycobactéries atypiques,
• aquariums,
• lampes UV

Abstract

Water is a natural habitat of mycobacteria. In aquaria 3 species of atypical mycobacteria are frequently present : M. marinera, M. kansasitand M. fortuitum. They are potential pathogen for fishes and men. Tuberculosis has been recognized as the cause of mortality in marine and fresh water fishes. Clinical signs of fish tuberculosis are variable : ascites, skin ulcerations, skeletal deformities. The human infection is cutaneous granuloma occuring after in jury in aquaria.

In the aquaria, of which two patients with cutaneous lesions due to M. marinum were analysed, UV lamps were not used. Many factors have an influence on the number of mycobacterial organisms in aquaria waters : number of fishes per tank, decontamination system, salinity, temperature.

To determine the consequence of each factor, a study has been conducted at the “Aquarium du Musée de Zoologie de Nancy”. Research of mycobacteria was carried out in the water of 40 tanks : 11 were supplied with fresh water and 29 with salt water. Each tank was equipped with germicide UV (λ : 253,7 nm) : the intensity was 15 watts for aquaria smaller than 1 000 liters and 36 watts for aquaria larger than 5 000 liters. The effectiveness of UV radiation against M. marinum was tested in 3 experimental fresh water tanks of 280 liters. The first part of the experiment was tested without fish. Tank n° 1 was a control, lamp was switched on during the complete study, M. marinum was not added. In tank n° 2 (with UV) and n° 3 (without UV), 2 ml of M. marinum (of suspension 10⁷ CF/ml) was added. Samples of water were analysed every two weeks. After six weeks tanks n° 2 and n° 3, were prepared for the next study : UV lamps were switched on in n° 2 and switched off in n° 3 both of which were contaminated by M. marinum. After 4 weeks 27 fishes, Cichlids, were introduced in the three aquaria. The day after, M. marinum was added to tanks n° 2 and n° 3. Every week water analysis was done, as well as an identification and quantification of all species of mycobacteria.

From each tank 250 ml of water were collected. The water was passed through a 0,2 mµ membrane. The filters were introduced in distilled water and decontaminated by lauryl sulfate. The culture of mycobacteria was grown with Loewenstein medium at 30 and 37 °C. Each colony type was identified by cultural and biochemical characteristics.

This study shows the richness in aquaria of mycobacteria; whatever the tanks, mycobacteria presence was constant. In non-treated home aquaria, the presence of mycobacteria was very important, 4 to 6 species per tank, (but in this case M. marinum was not found). In aquaria with UV lamps, the number of species per tank was lower (1 to 3).

The growth of mycobacleria could be prevented when the samples were contaminated by fungi and bacteria. However, inability to recover mycobacteria from water occurred only when a massive over-growth by non-mycobacterial contaminant was present (10³ CFU/ml). This was the case of non-treated tanks, belonging to patients who developed a chronic granuloma on their hands, M. marinum was not isolated in these aquaria. The evaluation of slowly growing mycobacteria could be altered by the important development of fast growing mycobacteria on the same culture tube. Among isolated species, M. fortuitum and M. gordonae saprophytic strains were frequent; M. kansasii and M. marinum involved in human cutaneous granuloma were unusual, as were the non-pigmented strains of groupe III of Runyon ; M. avium was not isolated.

During this study, we observed a relationship between the mycobacteria presence and the cleanness of tank and the fishes population. A great number of fishes per tank was a factor which increased the bacterial and mycobacterial contamination. During this experiment fishes didn't present tubercular-lesions but when a dead fish was examinated, the culture from post-modem samples revealed the presence of M. marinum. The microbiological examination of skip and viscera was negative.

The comparison of results in non-treated home tanks and UV treated tanks of the Museum indicates the role of water treatment by UV lamps on the number of isolated mycobacteria.

The germicide UV camps are frequently used for the decontamination of tanks. The efficiency is good for bacteria, but unknown for mycobacteria. This study shows that UV radiation decreased the mycobacterial contamination. The species of mycobacteria differ in their sensitivity to UV radiation. In experimental tanks, the results showed the great susceptibility of M. marinum to UV lamps such they were used in aquaria. Presence of fish dues not change the results. If the addition of M. marinum and the lighting of lamps were simultaneous, M. marinum was not isolated in water. If the contamination by M. marinum preceded the lighting of UV lamps, must of bacteria was eliminated in one week and the totality in 4 weeks. For the other species, we observed that the mycobacterial sensitivity to UV light decreases in the following order : quickly growing mycobacteria, photochromogen and scotochromogen strains. During our experimental study, M. gordonae was isolated more frequently when UV lamps were switched on. The results obtained in the 40 tanks with UV lamps allowed the evaluation of the influence of salinity and temperature of water on mycobacterial survival and the selection of species. We did not observed a difference in the concentration of mycobacteria in two types of aquaria, fresh water and salt water. Na Cl is known as an inhibitor of the mycobacterial growth. The sensitivity of strains differs. The salinity of water appears to be a selection factor. M. forfuitum was isolated more frequently in salt water.

M. marinum was isolated only in salt water and M. kansasii in fresh water. These results are surprising, as these strains have about the same metabolism. The temperature of water can also be a selection factor for mycobacteria, but in our study the temperature was similar in each aquarium (25°-26°). In this study, we did not observe thermophile strains such as M. avium.

Aquarists must be informed of the aquarium contamination by atypical mycobacteria and their role in the evolution of skin lesions after injury of hands and arms. The use of germicide UV lamps improves the bacteriological quality of water.

Keywords:

• Atypical mycobacteria,
• aquaria,
• UV lamps