In situ measured elimination of Vibrio cholerae from brackish water Marı ´a Elena Martı ´nez Pe ´ rez, Miroslav Macek and Marı ´a Teresa Castro Galva ´n National Autonomous University of Mexico, Campus Iztacala, Tlalnepantla, Mexico Summary In situ elimination of fluorescently labelled Vibrio cholerae (FLB) was measured in two saline water bodies in Mexico: in a brackish water lagoon, Mecoaca ´n (Gulf of Mexico; State of Tabasco) and an athalassohaline lake, Alchichica (State of Puebla). Disappearance rates of fluorescently labelled V. cholera O1 showed that they were eliminated from the environment at an average rate of 32% and 63 %/day, respectively (based on the bacterial standing stocks). The indirect immunofluorescence method confirmed the presence of V. cholerae O1 in the lagoon. However, the elimination of FLB was not directly related either to the presence or absence of the bacterium in the water body or to the phytoplankton concentration. keywords Vibrio cholerae, protozoan feeding, brackish water Introduction The position of Vibrio cholerae in the aquatic food web still attracts the interest of epidemiologists. This is not surprising taking into account the fact that among bacteria, typical for both plankton and sediment, the genus Vibrio prevails in the sea (Noble et al. 1990). Moreover, vibrios are supposed to be typical bacteria connected with an estuarine plankton – benthic annual cycle (Kaneko & Colwell 1975; Tamplin et al. 1990; Vanoy et al. 1992), suggesting the decomposition of chitin (particularly of crustaceans) as the natural source for this growth (Guthrie & Cofie 1991). During the last decades, V. cholerae has been reported from brackish or saline waters, mostly in a viable but non- culturable (VBNC) form (Xu et al. 1984; Colwell et al. 1985; Brayton et al. 1987; Huq et al. 1990; Tamplin et al. 1990; Corte ´s-Mun ˜ os et al. 2000). Bacterial cells with positive immune reaction, e.g. with agglutination serum, were visualized using fluorescent antibodies. The viability of the cells had been proven using a nalidixic acid test by Kogure et al. (1979), but this step was omitted (compare Brayton et al. 1987 with Huq et al. 1990). Although, generally, the apparently self-contradictory expression ‘viable-but-nonculturable’ has been applied to cells with various and often poorly defined physiological attributes (Bogosian et al. 2000), we should accept that direct counting using, for example, immunofluorescence tech- niques, gave us more realistic counts of bacteria such as V. cholerae in the environment. The microbial loop within a plankton food web (Azam et al. 1983) logically involves the niche of potentially pathogenic vibrios (V. cholerae included). Mechanical selectivity has been suggested as a major mechanism controlling suspension feeding of most of the bacterivorous organisms including protists, limiting the discrimination between particles entirely by the shape and volume, or by surface properties of a prey (Fenchel 1986). Suitability of different living bacterial prey to ciliate taxa has frequently been studied under laboratory conditions and V. cholerae (syn. V. comma) did not provide monoxenic growth (Dive 1973), while other vibrios might serve as an excellent source of food (e.g. Berk et al. 1976) or pathogenic vibrios might be ingested along with a suitable prey bacteria (e.g. Macek et al. 1997). In a natural environment, however, pathogenic or potentially pathogenic bacteria very quickly lose biochemical abilities, such as producing pigments and/or toxins. Whether the biochemical changes are also reflected in suitability of the bacteria for protozoa feeding remains unclear. For correct measurement of feeding rates, an offered prey must mimic the assemblage volume distribution and/ or must be of the same nature as the chosen food source studied (e.g. S ˇ imek et al. 1995). This is easily managed in the case of V. cholerae, which is prepared in concentrated nutrient medium (as this bacterium appears in nature during an epidemy) and labelled according to the protocol of fluorescently labelled prey (Sherr & Sherr 1993). Our aim was to analyse direct counts of V. cholerae using an immunofluorescence method and a potential Tropical Medicine and International Health volume 9 no 1 pp 133–140 january 2004 ª 2004 Blackwell Publishing Ltd 133