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A QUANTITATIVE MORPHOLOGICAL AND ECOLOGICAL STUDY OF PROTOZOA POLLUTING TAP WATER IN MEXICO ClTY F. RIVERA,A. ORTEGA, E. LÓPEZ-OCHOTERENA, and M. E. PAZ Protozoology Section, Department of Histology, Faculty of Medicine, National Autonomous University of Mexico, University City, Mexico 20, D.F. Mexico, and Laboratory of Protozoology, Faculty of Sciences, National Autonomous University of Mexico, University City, Mexico 20, D.F. Mexico RIVERA, F., ORTEGA, A., LÓPEZ-OCHOTERENA, E. & PAZ,M. E. 1979. A quantitative morphological and ecological study of protozoa polluting tap water in Mexico City. Trans. Amer. Micros. Soc., 98: 465469. The stiidy was performed using samples obtained from the tap water of the most populated area of the Federal District in Mexico City. The organisms were concentrated through filtration procedures and subsequently cultured in sterile wheat infusions. The cultures were obsemed for a week and the taxonomic identification was achieved to the leve1 of genus and species. Most commonly found were flagellates (84.58%), followed by amoebae (13.38%) and ciliates (1.86%). The greatest number (17) of species were found in the eastern region of the Federal District, though 16 were taken from the central region. Richest in numbers of organisms per m1 were Bodo edux (8636/m1), cysts of flagellates and ciliates (4484/m1), cysts of amoebae of the Vahlkampfiidae (2083/m1), and trophozoites of the genus Vahlkanapfru itself (14081ml). Among detected species, only those belonging to the Vahlkampfiidae and Hartmannellidae might have pathogenic effect on human beings. However, several species were found red which denotes a high degree of organic pollution in the supposedly potable waters of the Federal District. The fresh-water free-living protozoa have a cosmopolitan distribution as has been pointed out by Bovee (1957), Cairns (1962), Corliss (1973), and others. It is also well known that the diversity, density, and kind of protozoa present in waters may be used as indicators of their quality and potability (Cairns & Dickson, 1972). A widely used method to determine pollution is one employed in Europe named "Saprobic System" (Kolwitz & Marsson, 1908; see also Cairns et al., 1972). Therefore, since protozoa can pollute the drinkable waters which flow through the distribution duct system of a city, a study on this matter is well justified, especially in countries siich as Mexico where the knowledge on ecology of free-living protozoa is scant and mostly superficial. MATERIALS AND METHODS Some 25 random s a m ~ l e sof one gallon each were obtained directlv from the faucets of private hiuses receiviGg water from the distribution duit system that supplies the northern-eastern areas of the Federal District in Mexico City. Next, 95-liter water samples were filtered through an Ottawa sand column of 4 cm length and 1.5 cm interna1 diameter. A screen of cotton mesh was placed over the bottom and held in place with a hose clamp. The concentrated protozoa were freed from the sand after it had been removed from the column and thoroughly washed with distilled water. Then the organisms were in- Sample Total number of organisms per sample Percent Temperature (C) DH Bodo edax Klebs Bodo saltans Ehrenberg Euglena viridis Ehrenberg Anisonema truncatum Stein Anisonema emar~inatumStokes Ochromonas ~ y g s o t z k i unidentified species Trentonia flagellata Stokes Paranema trichophorum (Ehrenberg) Acanthamoeba astronyxis Ray & Hayes Acanthamoeba castellanii Douglas Amoeba verrucosa Ehrenberg Flabellula mira Schaeffer Hartmannella exundans Page Naegleria gruberi (Schardinger) trophozoite stage flagellate stage Vahlkampfia vahlkampfi Chatton Colpidium colpoda (Ehrenberg) Colpoda cucullus O. F. Müller Colpoda steini Maupas Sathrophilus agitatus (Stokes) Stylonychia pustulata Ehrenberg Tetrahymena pyriformis (Ehrenberg) Chilodonella cucullulus O. F. Müller Chilodontopsis vorax (Stokes) Platyophrya lata Kahl Organisms 1 2 2 3 4 5 2 546 97 6 5 690 560 7 5 8 547 9 TABLE 1 Incidence per m1 of organisms detected in 25 samples from Federal District 10 12 11 12 13 4,546 14 Sample Bodo edax Klebs Bodo saltans Ehrenberg Euglena ijiridis Ehrenberg Anisonema truncatum Stein Anisonema emarginatum Stokes Ochromonas Wyssotzki unidentified species Trentonia flagellata Stokes Paranema trichophorum (Ehrenberg) Acanthamoeba astronyxis Ray & Hayes Acanthamoeba castellanii Douglas Amoeba verrucosa Ehrenberg Flabellula mira Schaeffer Hartmannella exundans Page Naegleria gruberi (Schardinger) trophozoite stage flagellate stage Vahlkampjia vahlkampji Chatton Colpidium colpoda (Ehrenberg) Colpoda cucullus O. F . Müller Colpoda steini Maupas Sathrophilus agitatus (Stokes) Stylonychia pustulata Ehrenberg Tetrahymena pyriformis (Ehrenberg) Chilodonella cucullulus O. F . Müller Chilodontopsis voraz (Stokes) Platyophrya lata Kahl Total number of organisms per sample Percent Temperature (C) PH Organisms 15 16 17 18 19 TABLE 1 Continued 20 21 22 23 24 25 Total Percent 468 TRANS. AMER. MICROS. SOC., VOL. 98, NO. 3, JULY 1979 oculated into sterile wheat infusions, covered with sterile cotton caps, and incubated at roorn temperature (15-25 C) for a period of seven days. Identification and counting of isolates of each culture were performed daily, using a photornicroscope (phase contrast, 40 x and 100 X ) and a Neubauer counting chamber. Identification and incidence of the concentrated speciinens per ml, as well as such variables as temperature and pH from the 25 tap water samples are given in Table 1. Only the trophozoite stage and the active form of the organisms observed are considered in this table, although we have also found the encysted stage of some ciliates, flagellates, and arnoebae as well as some palmella stages of phytoflagellates. Total numbers of organisms and percentages of species are also included in the table. The most polluted tap waters were those from the eastern-western sides of the explored area of the Federal District, followed successively by northern, central, and southern sides. The most comrnonly found protozoa were flagellates, followed by amoebae and ciliates. Among the flagellates, the most abundant forms were Bodo edax, B. saltans, and Ochromonas sp. Within the amoeboid group, most frequently found were members of the Vahlkampfiidae family (trophozoic as well as cystic stages), mainly of the genus Vahlkampjia itself. Among ciliates, most abundant were Colpidium colpoda, Colpoda steini, and Tetrahymena pyriformis. This is the first research ever carried out in Mexico concerned with protozoa that may act as pollutants or as indicators of nonpotability of tap water. The results clearly demonstrate that the theoretically drinkable waters of the Federal District contain protozoa, either in their trophozoic form or cystic stage. Some of the detected organisms belong to genera that may be potentially pathogeriic when inoculated into experimental animals, viz., Naegleria and Acanthamoeba. We also discovered organisms that are widely considered to be indicators of pollution in drinkable waters. For example, we found such saprobic organisms as Euglenu viridis, Colpidium colpoda, Sathrophilus agitatus; and such mesosaprobic forms as Bodo edax, B. saltans, and Colpoda cucullus. Another interesting finding was that some of the sainples examined did not show any kind of protozoa at any time during our observations, only bacteria (see Table 1). Still other samples showed only cystic stages of flagellates, amoebae. or ciliates. Some óf the microorganisms found have not been reported before in Mexico (i.e., they are not listed by López-Ochoterena & Roure-Cane, 1970). These include: Anisonema truncatum, Chilodontopsis vorax, Flabellula mira, Hartmannella exundans, Sathrophilus agitatus, and Trentonia jiagellata. The bacteriophagous protozoa detected in the samples, which may be considered as decontaminating organisms since they capture bacteria, include Ochromonas sp., Peranema trichophorum, Acanthamoeba astronyxis, A. castellanii, Amoeba verrucosa, Flabellula mira, Hartmannella exundans, Naegleria gruberi, Vahlkampjia vahlkampji, Colpidiutn colpoda, Tetrahymena pyriformis, Chilodonella cucullulus. BOVEE,E. C. 1957. A small amoeba of fresh water lakes and ponds, Majorella cultura. Tran-s. Amer. Micros. Soc., 80: 54-62. CAIRNS,J., JR. 1962. The environmental requirements of fresh-water protozoa. Zn Biological Problems in Water Pollution, 3rd Serninar, August 1962, PHS Publ. No. 999-WP-25, pp. 48-52. CAIRNS,J., JR. & DICKSON,K. L. 1972. An ecosystematic study in the South River, Virginia. Water Res. Ctr., Virginia Polytechnic Institute, Blacksburg, Virginia, Bull. 54, pp. 1-104. CAIRNS,J., JR., LANZA,G. R. & PARKER,B. C. 1972. Pollution related structural and functional changes in aquatic communities with einphasis on freshwater algae and protozoa. Proc. Acad. Nat. Sci., Philadelphia, 124: 79-124. CORLISS,J. 0. 1973. Protozoan ecology: a note on its current status. Amer. Zool., 13: 143-148. KOLWITZ,R. & MARSSON,M. 1908. 0kologie der pflanzlichen Saprobien. Ber. Deutsch. Bot. Ges., 26: 509-519. L~PEZ-OCHOTERENA, E. & ROURE-CANE,M. T. 1970. Lista taxonómica comentada d e protozoario~d e vida libre d e México. Rev. Soc. Mex. Hist. Nat., 31: 23-68.