Amoebae in a waste stabilization pond system in Mexico

l

AMOEBAE IN A W ASTE ST ABILIZATION POND SYSTEM IN

MEXICO

FERMIN RIVERA 1 *, GISELA GARCIA 2, ALFONSO LUGO 1

,

ELENA ZIEROLD 1, JESUS ISLAS 1

, ELIZABETH RAMIREZ 1,

and PATRICIA BONILLA 1

1 Project of Conservation and lmprovement of the Environment, Unit of Interdisciplinary Research of Hea/th and Education Sciences (U.I.I.C.S.E.) E .N.E .P. IZTACALA, U.N.A.M. , Los Reyes Iztacala, Estado de

México, México 2 General Direction of Ecological Order and Environmental Impact, Subsecretary of Ecology, Secretary of

Urban Development and Ecology, México , D.F.

(Received April 22, 1985; revised August 29, 1985)

Abstract. A protozoo1ogical survey was performed to analyze the organisms of the subphylum Sarcodina Schmarda, present in waste stabilization ponds located at Santo Tomás Atzingo, México, from March to December 1981. The amoebae isolated were identified and counted. Several physicochemical parameters were also determined and correlated with the biological data.

Thirteen amoebic species were isolated and identified: one definite pathogen, Entamoeba histolytica Schaudinn; three species that have shown pathogenic strains for humans: Acanthamoeba polyphaga Page, A canthamoeba castellanii Douglas, and N aegleria sp. (Alexeieff) Calkins; two opportunist species : Entamoeba coli (Grassi) Casagrandi and Barbagallo, and Iodamoeba bütschlii (Von Prowazek) Dobell ; and seven free-living amoebae: Acanthamoeba astronyxis Ray and Rayes, Amoeba proteus (Pallas) Leidy, Hartmannella exundans Page, Hartmannella vermiformis Page, Pelomyxa palustris Leidy, Vahlkampfia avara Page, and Vahlkampfia russelli Singh.

A clear correlation between the number ofamoebae and the temperature and alkalinity ofthe system was found. The removal capacity of the ponds for E. histolytica varied from 30 to 100 % during the survey. The amoebae showed a succession in space and time in the system studied.

l. Introduction

There has been an increase in water pollution in the recent years due mainly to population growth and industrial development; this has made wastewater treatment necessary in many communities and in various industries. A stabilization pond offers a good example of a simple and an inexpensive form of wastewater treatment for a community especially in many developing countries. Stabilization ponds are considered to be secondary biological treatment systems, in which biological conversion of the organic matter is carried out by various organisms to produce new living matter.

The role of protozoa in wastewater treatment systems is debated (Hanel, 1979). However, since most protozoa are bacteriophagous (Grassé, 1952) it is supposed that they may actas bacterium-reducing organisms. Furthermore, bacteriophagous amoebae, together with other kinds of phagocytic protozoa, constitute links in the transfer of energy through the trophic levels of the food chain (Sleigh, 1979).

There are al so pathogenic amoebae such as E. histolytica and many of the Naegleria

Acanthamoeba group, the presence of which, in the effiuent of treatment systems, may

* Author for all correspondence: Av. Universidad # 220-4, México 03020, D.F., México.

Water, Air, and Soil Pollution 28 (1986) 185-198. © 1986 by D. Reidel Publishing Company.

186 FERMIN RIVERA ET AL.

be hazardous to health. The pond system analyzed in this study is located close to a small village [Santo Tomás Atzingo] of the state of Mexico at 19° 12 ' 10" north and 98 o 48 ' 00" west longitud e, at 24 7 5 m abo ve sea level. The prevailing clima te of the are a

Lagoon 2 Lagoon 1

ll A

III I

3

.___;~-IV ~ I I

---r__r--

1 1

l 1

llr l ·1 E

---14.6 ...,m..._ __ +----14. 6 .... m-.--__

Fig. l. The waste stabilization pond system at Sto. Tomás Atzingo, Showing the sampling stations. Key of stations : A: lnfluent

1: Center 1 11 : Center 2 111 : Center 3 IV : Center 4 E: Effiuent

\

\

1

\

\

1

AMOEBAE IN A W ASTE ST ABILIZA TION POND SYSTEM IN MEXICO 187

is temperate subhumid, with summer rains. The mean annual temperature is 14.1 oC with a maximum of 29 oC and mínimum of - 3 o C. The pond system receives the wastewater from the small village with 1200 people who are dependent on agriculture and cattle raising. The pond system dimensions are given in Figure l.

The aim of the present survey was to isolate, identify, classify and quantify the amoebic specimens present in the ponds, and also to relate the species found with sorne physicochemical parameters determined for such waters.

2. Material and Methods

2.1. FIELD WORK

2.1.1. Sampling

Six stations were selected (Figure 1 ). Twenty samples were taken, two per month, from March to December 1981.

Surface samples were recovered using sterilized 500 mL glass bottles. Samples from the bottom were taken with the use of a 2 L Van Dorn bottle. These samples were used to observe the protozoa in vivo. Another set of 100 mL samples were also recovered in 125 mL bottles, and fixed after collection with 1 mL of 4% formaldehyde. These fixed samples were used to quantify the organisms.

2.1.2. Physicochemical Parameters

Samples were also taken and processed in situ for the determination of the following parameters: dissolved oxygen (DO), analyzed by the Winkler method with azide modification as in 421 B in Standard Methods (1980); pH, potentiometrically measured as in 423 in Standard Methods (1980); C02 , analyzed by the titrimetric method as in 406 B in Standard Methods (1980); temperature, taken by the direct method as in 212 in Standard Methods (1980); acidity, as in 402 in Standard Methods (1980); hardness, by the EDTA titrimetric method as in 314 B in Standard Methods (1980); and alkalinity, as in 403 in Standard Methods (1980).

2.2. LABORATORY WORK

2.2.1. Culture Media

(a) Chalkley's medium with rice grains (Mackinnon and Hawes, 1961), for free-living amoebae incubated at 20 o C.

(b) Balamuth and Sandza medium (Kudo, 1976), for Entamoeba histolytica. Blanks were prepared for each culture medium. Observation of inoculated culture media and blanks was performed every three days by light microscopy, phase contrast - 10, 40,

and 100 x, for two weeks.


2.2.2. Protozoa Counting

An estimation on the incidence of each amoebic species was made from May to December 1981, using the Neubauer chamber (Singleton and Sains, 1981) for the last eight samples.

2.2.3. Fixation and Staining

The Bouin fluid was used for fixation and carmine borax to stain the smears (Gaviño et al., 1977). The Heidenhain's iron hematoxylin was also used to make permanent smears (Mackinnon and Hawes, 1961 ).

2.2.4. Identification

The morphological criteria followed for the identification of the amoebae were those suggested by Grell (1973), Jahn et al. (1979), Kudo (1976), Page (1966), Mackinnon and Hawes (1961) and Westphal (1978).

2.2.5. Physicochemical Analyses

The following parameters were measured in the laboratory: 5 da y biochemical oxygen demand [BOD5 ] analyzed as in 507 in Standard Methods (1980); ammonia N, as in 417 B in Standard Methods (1980); nitrites as in 419 in Standard Methods (1980); nitrates, as in 418 e in Standard Methods (1980); total residue, as in 209 A in Standard Methods (1980); total suspended matter, as in 209 D in Standard Methods (1980); phosphates, as in 424 E in Standard Methods (1980); and sulfates as in 426 e in Standard Methods (1980).

2.3. STATISTICAL ANALYSIS

The correlation coefficient of Pearson was used (Sokal, 1978), and it was observed that for a 0.5 value of the coefficient there was a formation of natural groups of the physicochemical parameters.

For cluster analysis the complete linkage clustering method was chosen (Sokal and Rohlt, 1980).

3. Results

The thirteen amoebic species isolated from the system are indicated in Table I. Among the species isolated E. histolytica is a definite pathogen; A. polyphaga, Naegleria sp., and A. castellanii have shown strains capable of producing eye infection (Nagington et al., 1974; Visvesvara et al., 1975) and granulomatous encephalitis (Martínez et al., 1977) in humans; and E. coli and /. bütschlii may become opportunist pathogens for humans (Faust et al., 1981). The other seven species found typically free-:-living, either bacteriophagous such as A. astronyxis, H. exundans, H. vermiformis, V. avara, and V. russelli (Sundstrom, 1979) or feeding on ciliates and tlagellates like A. proteus, or on algae like P. palustris (Kwang, 1973). The incidence ofthe species during the survey was variable but the greatest number (8 species) appeared on June and September in pond 1, and also

.•

TABLEI Amoebic species found in the waste stabilization pond system at Santo Tomas Atzingo from March to December 1981

Species Mar Apr M ay Jun Jul Aug Sep Oct Nov

Acanthamoeba astronyxis Ray and Hayes X X X Acanthamoeba caste/lanii Douglas X X X X X xo o Acanthamoeba polyphaga Page X X X Amoeba proteus (Pallas) Leidy X X X X Entamoeba coli (Grassi) Casagrandi and Barbagallo X X Entamoeba histolytica Schaudinn X X X X X xo xo Hartmannella exundans Page X X X X xo X Hartmanne/la vermiformis Page X X xo xo Iodamoeba bütschlii (Von Prowazek) Dobell X X Naegleria sp. (Alexeieft) Calkins X X X X X xo Pelomyxa palustris Leidy X X X Vahlkampfia avara Page X o V ahlkampfia russelli Singh X X X X

X = Species found in pond 1 O = Species found in pond 2 1 = lncidence of amoebae in pond 1 (%)

De e 1

30% xo 80%

30% xo 50%

20% xo 80% X 70%

40% 20%

xo 70% 30% 20%

X 50%

> ~ o tTl l:!l > tTl

z > ~ > Cl'l ..., tTl Cl'l ..., > l:!l ¡:::: Ñ > ..., o z "C o z t:l Cl'l ....:: Cl'l ..., tTl ~

z ~ tTl ~ ñ o

00 1,0

TABLE II

Number of amoebae isolated per mililiter and per month from March to December, 1981 in pond 1 ~ o

Species Depth Mar Apr M ay Jun Jul Aug Sep Oct Nov De e

Amoeba proteus S 80 30 80 o o 60 o o o 20 B o 20 o o o o o o o 40

Acanthamoeba astronyxis S o o o 120 o 90 50 o o o B o o o o o 60 20 o o o

Acanthamoeba castellanii S 20 530 120 90 70 o 20 o o o B o o o 80 80 o 140 o o 270

Acanthamoeba polyphaga S o o o o 30 o 340 o 420 o B o o o o o o 600 o 60 o

Entamoeba coli S o 60 o o o o o o o o "rJ

B o o o o o o 20 o o o tT1 :;el ~

Entamoeba histolytica S o 2980' 470 1880 1210 990 20 o 20 130 z B o 2560 150 1760 2160 540 80 o 80 250 :;el

< 1 odamoeba bütschlii S o o o 20 o o o o o o tT1

:;el

B o o 60 10 o o o o ;¡.

o o tT1 ..., H artmannella exundans S o o 470 80 o o 210 o o 40 ;¡.

r B o 280 o o o 100 170 o 700 60

H artmannella vermiformis S o o 80 o 50 o 410 50 o o B o o 110 o 950 o 30 130 o o

Naegleria sp. S 250 380 600 3770 o o 100 230 o 200 B o 90 o o o o 90 660 o 510

Pelomyxa palustris S o o o 40 o 50 o o o o B o o o o o 20 o o o o

Vahlkampfia avara S o o o 750 o o o o o o B o o o o o o o o o o

V ahlkampfia russelli S 400 50 150 130 o o o o o 110 B 450 300 1000 2500 o o o o o 100

S: Surface B: Bottom ....

~

,.)

AMOEBAE IN A WASTE STABILIZATION POND SYSTEM IN MEXICO 191

TABLE III

Number of amoebae isolated per mililiter and per month from September to December, 1981 in pond 2

Species Depth* Sep Oct Nov De e

Amoeba Proteus S o o o 10 B o o o o

Acanthamoeba castellanii S 30 o 40 120 B o o 150 110

Acanthamoeba polyphaga S o o 70 o B o o 20 o

Entamoeba histolytica S 10 o 10 80 B o o o 110

H artmannel/a exundans S 20 o o o B o o o o

H artmannella vermiformis S 30 o o o B o 150 o o

Naegleria sp. S o o o 50 B o 150 o 20

Vahlkampfia avara S 220 o o o B o o o o

*S: surface B: bottom

on September ( 5 species) in pond 2. On the other hand the lowest number ( 4 species) was found on March in pond 1, and on October (one species) in pond 2. The results of the quantitative analysis are shown in Tables II and III. The most abundant species were E. histolytica found almost always in the cystic stage and on the bottom of the ponds; Naegleria sp., H. exundans, H. vermiformis, and V. russelli were found only in the trophic stage and in the surface strata of the ponds. In general, more amoebae were isolated from the bottom than from the surface of the ponds. Al so, trophic forms were found in the surface and cysts on the bottom of the system. The physicochemical parameters determined and their values are indicated in T ables IV and V. It was observed that an increase ofthe C02 concentration in pond 1 and 2 correlated well with a higher incidence ofthe free-living and the amphizoic amoebae especially Naegleria sp., H. exundans, andA. castellanii. In contrast, high levels of C02 in the system correlated with low numbers of specimens of E. histolytica, while low C02 corresponded with an increase ofthe number of amoebae ofthis species. The same pattern was shown by the alkalinity of the system in relation to the incidence of amoebae.

The efficiency of the system for removing amoebae was variable along the survey and in relation with the species considered. Thus, A. castellanii was removed only in November (20 %) and December (50%); H. exundans in June (100 %) and September (50%); E. histolytica in April (100%), May (100%), June (100%), August (57%), September (100 %) and November (30 %); Naegleria sp. in June (60 %) and September

~ TABLEIV N

Mean values of the physicochemical parameters determined from March to December 1981 in pond 1

Parameter Depth Mar Apr M ay Jun Jul Aug Sep Oct Nov De e

Transparency in cm (Secchi disc.) 4.5 5.5 4.5 5.2 4.2 4.0 4.5 4.2 5.1 5.7

Temperature oc S 21.8 19.2 17.7 19.3 20.8 20.6 19.9 18.8 14.6 13.5 B 20.0 20.0 21.0 19.5 17.7 18.2 19.8 17.5 13.7 14.5

pH S 6.2 7.1 7.1 7.3 7.5 7.5 7.6 7.6 7.6 8.0 B 6.1 7.2 7.2 7.1 7.1 7.5 7.5 7.2 7.5 7.5

C02 (mg L - 1) S 36.1 34.9 35.2 57.6 37.2 34.2 37.7 99.3 61.1 54.1

B 37.2 33.8 34.9 56.3 35.7 30.2 36.1 93.8 66.5 46.3 'Tj m

Alkalinity (mg L - 1) S 305.0 306.1 302.5 304.3 243.0 255.9 498.1 271.6 357.8 392.6

:;o 3::

B 266.0 272.0 265.4 302.5 239.7 277.5 382.0 332.2 103.0 285.0 z :;o

Hardness (CaC03 ) (mg L - 1) S 44.0 42.0 45.0 31.6 48.3 56.6 28.3 115.0 173.3 100.0 ~ m

B 23.0 24.0 25.0 27.5 57.5 42.5 27.5 85.0 232.5 125.0 :;o ;¡.. m

Acidity (CaC03 ) (mg L - 1) 83.6 S 81.2 25.1 33.2 35.0 93.6 93.6 306.4 145.6 125.8 >-l

B 85.3 88.1 33.6 27.7 41.1 105.4 90.8 414.2 156.0 121.0 ;¡.. r

Dissolved Oxygen (mg L - 1) S o o o o o o o o o o

B o o o o o o o o o o N-NH3 (ppm) S 25.0 23.8 24.6 26.0 35.2 38.6 51.8 68.1 43.8 50.2

B 30.1 29.6 31.2 33.8 35.4 27.0 26.2 35.0 29.9 32.3

N-N02 (ppm) S 0.34 0.36 0.35 0.38 0.45 0.45 0.56 0.63 0.53 0.60 B 0.42 0.43 0.41 0.47 0.56 0.58 0.84 0.79 0.52 0.51

N-N03 (ppm) S 0.92 0.89 0.18 1.02 0.15 0.12 0.18 0.62 1.39 1.29 B 0.91 0.88 0.86 0.87 0.12 0.14 0.20 0.66 1.06 1.07

Depth: S = Surface B = Bottom

,._.


TABLEV

Mean values of the physicochemical parameters determined from September to December 1981 in pond 2

Parameter Depth* Sep Oct Nov De e

Transparency in cm (Secchi disc) 5.25 5.0 6.0 5.7

Temperature oC S 21.1 21.1 14.8 13.6 B 16.5 17.5 13.8 14.3

pH S 7.2 7.2 7.7 7.5 B 7.7 7.2 7.8 6.8

C02 (mgL - 1) S 37.6 33.9 70.6 56.6

B 41.9 34.9 60.4 68.8

Alkalinity (mg L - 1) S 357.9 358.7 174.1 281.8

B 375.2 299.0 90.2 294.5

Hardness (CaC03 ) (mg L - 1) S 70.6 98.3 216.6 116.6

B 97.7 92.5 275 .0 132.2

Acidity (CaC03 ) (mg L - 1) S 113.50 287.53 155.11 132.2

B 181.60 419.95 139.03 66.5

DO (ppm) S o o o o B o o o o

N-NH3 (ppm) S 30.4 21.4 24.8 23.4 B 11.2 23.8 27.0 25.2

N-N02 (ppm) S 0.59 0.41 0.76 0.68 B 0.95 0.92 o 0.30

N-N03 (ppm) S 0.10 0.65 0.90 0.80 B 0.12 0.62 0.58 0.52

*S : surface B: bottom

(84% ). The other nine species were never isolated from the eflluent of the system so one can assume that their removal was complete (100 %) all the time.

Among the amphizoic amoebae only A. castellanii was detected (in May, June, July, September, November and December) in the influent of the system; and among the free-living species only H. exundans was found (May) in the influent of the system. E. histolytica was isolated all the time along the survey at the influent station.

The cluster analysis of the physicochemical parameters showed the existence of three groups (Figure 2). The first one was represented by water temperature which exhibited a proportional effect over the number of amoebae in the system during the concentration period.

The second group was formed by the alkalinity. In September 1981 the alkalinity showed a remarkable increase associated with a change ofthe water color in the ponds. The water color varied from brown to green. This change of the alkalinity values may be considered as an indicator of the variation of the productivity of the system (Cole,

194

1: o

Water

Tempera tu re

Al kal i nity

pH

Aci dity

Total

Ha rdness

Free C02

FERMIN RIVERA ET AL.

Pearson's Correlation Coefficients

0:5 l

-

r-

Fig. 2. Dendogram prepared by the complete linkage clustering method showing correlations among physicochemical parameters.

1975). Furthermore, the alkalinity may be increased by the liberation from the anoxic sediment of Fe and Mn bicarbonates, and by the anaerobic production of ammonia bicarbonate by bacteria (Wetzel, 197 5). This fact may explain why the alkalinity appears non related - through the cluster analysis - to such parameters as pH, acidity and free C02 , variables to which it is normally related. The existence of a wide anaerobic zone el ose to the bottom of the ponds favors the production and liberation of high amounts of bicarbonates. Thus, the alkalinity of the ponds is due almost exclusively to the bicarbonates.

The third group was composed of pH, acidity, total hardness and free C02 . This group was related with physicochemical changes of the ponds caused by the processes of dilution and concentration. pH levels showed a great dependence on concentration and dilution periods. The degradation of organic matter, a result of the metabolism of anaerobic bacteria resulted in an increase of free C02 and acidity levels. This phenomenon was aspecially evident during N ovember 1981.

4. Discussion

The pond system studied performed anaerobically, mainly because of its high concentration of organic matter and low transparency. The absence of DO leads to the anaerobic formation of C02 , a fact that explains the high concentrations of this gas found in the system as well as the increased alkalinity. Alkalinity and temperature were

1

AMOEBAE IN A W ASTE ST ABILIZA TION POND SYSTEM IN MEXICO 195

the most predominant parameters in the performance of the system. Alkalinity correlated well with changes ofproductivity ofthe ponds. The slightly basic pH ofwater was a factor that favored the growth of amoebae (Smith, 1968).

Hardness of the system waters varied from moderate (Oto 150 mg L - 1 of CaC03 )

to in tense (more than 250 mg L - 1of CaC03 ) due perhaps to the Ca and Mg ions in the infiuent.

water temperature showed a range from 13.5 to 21.8 o e in pond 1 ; and from 13.6 to 21.1 oC in pond 2. Temperature was then a factor that favored the amoebic growth of free-living and amphizoic species (Hart, 1974), but was a limiting factor for the adequate growth of definite pathogens such as E. histolytica or the opportunistic ones such as E. coli and /. bütschlii, species which grow better at higher temperatures. This may have been one of the reasons to find the latter species mainly in the cystic stage while the free-living and the amphizoic amoebae were found preferably in the trophic stage.

The N in the system was found mainly in the form of ammonia, the formation ofwhich was favored by anaerobiosis. However, significant amounts of nitrates and nitrites also were found. This latter finding could be explained by the nitrifying processes carried on by bacteria in the surface strata of the ponds.

With respect to protozoological incidence thirteen amoebic species found in the lagoons belonged to the following orders, suborders and families (Page, 1966; Rivera and Paz, 1978; Levine et al., 1980): A. proteus, arder Amoebida, suborder Tubulina; A. astronyxis, A. castellanii andA. polyphaga, arder Amoebida, suborder Acanthopodina, family Acanthamoebidae; P. palustris, arder Pelobiontida; E. coli, E. histolytica, and /. bütschlii, arder Amoebida, suborder Tubulina, family Endamoebidae; V. avara,

V. russelli, and Naegleria sp. order Schizopyrenida, family Vahlkampfiidae; H. exundans

and H. vermiformis, arder Amoebida, suborder Acanthopodina, family Hartmannellidae.

DO was never detected with the methods used but there were reasons that made us suspect the presence of this gas at least in low levels such as the high incidence of phytofiagellates and microscopic algae in the ponds, the exposure of the pond waters to the natural aeration of the wind, the presence of strict aerobic amoebae such as A. proteus and P. palustris that need an optimal level of 10 ppm of DO to grow and reproduce (Kwang, 1973), and the formation of oxidized compounds like nitrates and nitrites that were present at high levels in the ponds. This means that the low levels of DO present in the ponds must have been used up immediately by aerobic organisms (BOD) or by compounds in process of oxidation (COD). However one must also suspect the existence of completely anaerobic spots in the lagoons, especially in the zone close to the bottom, where only the anaerobes (strict offacultative) can thrive (Lacy and Rivera, 1982; Carranco et al. , 1984).

These facts may explain well the presence of trophic amoebae preferably close to the surface strata of the ponds where the microalgae are found and the wind aerates the waters, and of the cystic amoebae which are found deeper or even at the bottom.

The efficiency ofthe system for removing amoebae varied much according to time and


species considered and it was found to remove E. histolytica (30 to 100% ), Naegleria sp. (60 to 84%), H. exundans (50 to 100%), andA. castellanii (20 to 50%).

Among the small amoebae all the species other than A. castellanii and H. exundans

were not isolated from the influent of the system, which means that they may have reached the pond waters either by air (Rivera et al., 1984) or directly from the soil of the ponds (Rivera et al., 1978, 1983).

All amoebae found in the ponds were well known bacteriophagues (Duncan, 1974). Because E. histolytica, E. coli and /. bütschlii are coprophilic, one can assume that they would be good grazers on the coliform bacteria so profusely present in the system; in this sense, the amoebae isolated may have been important consumers in the ponds, especially in June and September 1981 (Leadly, 1980).

As expected (James and E vi son, 1979) of the thirteen species found only Naegleria sp., A. castellanii, H. exundans, andE. histolytica tolerated well the extreme conditions of pollution of the lagoons and became therefore, the predominant species. It seemed clear that the survival and reproduction of amoebae depended, in the ponds studied, on the quality and composition of the influent and on the purifying capability of the system as a whole.

The influent also determined the periods of concentration and dilution in the ponds, affecting therefore the environment where the amoebae thrived.

The appearance of the four most abundant amoebic species in the system showed a clear pattern of incidence and succession.

At the beginning of the survey (March) Naegleria sp. was the most abundant species followed closely by A. castellanii. At that time E. histolytica and H. exundans hada lower incidence. Later on (April), the incidence of Naegleria sp. decreased slightly while that of A. castellanii and H. exundans increased. These two latter species became finally the most numerous, leaving E. histolytica and Naegleria sp. in the third and fourth ranks (May). On June Naegleria sp. and A. castellanii became the most abundant species whereas on July, August and September E. histolytica, H. exundans, andA. castellanii

were the most frequently found. On October Naegleria sp. became again the predominant species whereas on November H. exundans predominated in the ponds. Finally, on December Naegleria sp., E. histolytica, and A. castellanii became the most frequently found species.

The data in Table IV and V confirm the existence of polysaprobic conditions in the system (Sládecek, 1973). Because ofthe conditions the BOD5 ofthe waters decreased, so did the concentrations of total residue, suspended matter, faecal streptococci and coliforms.

An evaluation of the degree of organic pollution of the ponds, on the basis of the Sládecek system (1973) would reveal conditions varying from isosaprobity and mesosaprobity. This range is characterised by a predominance of decomposers over consumers and producers, as is confirmed by this survey.

·'


5. Summary

On the basis of the present findings we can conclude that the prevailing amoebic species of the pond system were free-living, amphizoic and al so bacteriophagous and coprophilic. The system performed anaerobically. The temperature of the waters favored the growth and proliferation of the free-living and amphizoic amoebae and was partially a limiting factor for amoebae of the Endamoebidae family. The same pattern was shown by the col concentration and the alkalinity of the system in relation with the incidence of amoebae.

On certain occasions the amoebae became important consumers. A change of incidence and succession of the four most abundant species was present in samples related to variable concentrations of organic matter present in the waters.

The efficiency of the system for removing amoebae was variable and related to the species considered. The elimination of E. histolytica was rather good ( 100%) most of the time; however, there were occasions in which this removal only reached 30%; therefore, it would be advisable to improve the efficiency of the system and al so to use the water of the effluent to irrigate crop lands with plants whose fruit will not be in direct contact with water.

Acknowledgments

This study was performed with the financia! support of CONACyT (National Council of Science and Technology). The English version ofthis manuscript was kindly revised by Skidmore Smith MacDonald, Language Section, ENEP-Iztacala, UNAM. Alma L. Padilla and Ma. Luisa Islas provided valuable secretaria! assistance.

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Amoebae in a waste stabilization pond system in Mexico

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