The Journal of Published by

the International
Eukaryotic Microbiology Society of
Protistologists

J. Eukaryot. Microbiol., 57(4), 2010 pp. 362–368

r 2010 The Author(s)
Journal compilation r 2010 by the International Society of Protistologists
DOI: 10.1111/j.1550-7408.2010.00484.x

A New Species of the Genus Metacystis (Ciliophora, Prostomatida, Metacystidae)

from a Wastewater Treatment Plant
LUCÍA ARREGUI,a BLANCA PÉREZ-UZ,a ANDRÉS ZORNOZAb and SUSANA SERRANOa
a
Departamento de Microbiologı´a III, Facultad de Biologı´a, Universidad Complutense, 28040 Madrid, Spain, and
b
Grupo Aguas de Valencia, EDAR Quart-Benager, EPSAR, Valencia, Spain

ABSTRACT. Unusual prostomatid specimens were found in the biological reactor of a wastewater treatment plant in a health resort in
Valencia, Spain. These ciliates were attached to flocs unlike other free-swimming prostomatid ciliates described to date in the mixed liquor
of activated sludge plants. The morphological study of this species led to a typically different combination of characteristics: elongated
cell shape, 20–30 somatic kineties, 2 perioral kineties, and 1 circumoral kinety, 1 large vacuole protruding at the terminal end, a lorica
tapered toward the aperture with a smooth neck, and 11–16 annular ridges. These characteristics place this representative as a new species
of the genus Metacystis—Metacystis galiani n. sp. This species became the dominant population within the biological reactor when high
values of conductivity (4,244 mS/cm) and temperature (26.8 1C) were recorded.
Key Words. Activated sludge, high conductivity habitat, protist.

MATERIALS AND METHODS

B IOLOGICAL reactors of sewage treatment plants receive in-
puts with different physical–chemical characteristics that de-
termine variable biocenosis structures. The main components of
Sessile ciliates were sampled from the biological reactor of a
WWTP of a health resort located in Cofrentes, Valencia, Spain
this ecosystem are bacteria that reach 90–95% of the biomass, (39113 0 N, 1104 0 W). Samples were collected in plastic bottles and
while protists and other microorganisms represent just 5–10% of taken to the laboratory for analysis. Fifty-microliter aliquots con-
the total biomass (Bitton 2005). The established community is taining living cells were observed with a phase contrast micro-
organized as bioaggregates or flocs in which the biotic component scope using either an Olympus BH-5 (Olympus Optical España
is embedded in a complex matrix composed of organic and inor- S. A., Madrid, Spain) or a Zeiss Axiostar microscope (VWR In-
ganic particles (contained in the sewage) and extracellular poly- ternational Eurolab S. L., Barcelona, Spain). Digital images were
meric substances produced by the microbial activity (Barker and taken with a digital camera (Olympus Camedia C-5050 or a Dig-
Stuckey 1999; Laspidou and Rittmann 2002). ital Still Camera Sony DKC-CM30) attached to the microscope.
Ciliates, often one of the most significant groups of protists Linear measurements were obtained from the digital images with
within this ecosystem, have been divided into two categories de- Adobe Photoshop 6.0 after calibration with a 1-mm Zeiss stage
pending on their ecological niche inside the reactor (Madoni micrometer.
1988; Martı́n-Cereceda, Serrano, and Guinea 1996, 2001; Poole Ciliates were stained using the pyridinated silver carbonate
1984): (i) associated with flocs are two functional groups—crawl- method of Fernández-Galiano (1976, 1994) and the protargol im-
ing ciliates with flattened bodies and a specialized ventral ciliature pregnation modification of Wilbert (1975). Permanent slides of
allowing the movement on the floc surface and sessile ciliates at- both staining techniques were deposited at the Museo Nacional de
tached to the floc through stalks or adhesive structures; and (ii) Ciencias Naturales (Madrid, Spain) with accession number
free-swimming ciliates moving freely in the mixed liquor. The MNCN 39.04/1.
first category of ciliates is generally more abundant (Curds and The specimens were also stained with Flutax-2, a fluorescent
Cockburn 1970; Madoni 1988; Martı́n-Cereceda, Serrano, and derivative of taxol that binds specifically to microtubules (pro-
Guinea 1996). vided by J.M. Andreu of the Centro de Investigaciones Biológicas,
Species of prostomatid ciliates are scarce in conventional ac- Consejo Superior de Investigaciones Cientı́ficas, Madrid) as de-
tivated sludge wastewater treatment plants (WWTP), never reach- scribed in Arregui et al. (2003) in order to acquire complementary
ing higher numbers or reported as the dominant population under data to those obtained with the silver impregnations since optimal
stable reactor conditions (Curds 1969; Esteban, Téllez, and Ba- silver impregnation images were difficult to obtain due to the
utista 1991; Madoni 1988; Martı́n-Cereceda, Serrano, and Guinea presence of a lorica. Samples were observed under a Zeiss Axio-
1996). Exceptionally the prostomatid swimming ciliate Coleps plan 2 epifluorescence microscope using 40X and 63X objectives.
hirtus has been described in biological reactors showing high effi- Images were recorded with a CCD Spot Camera and then ana-
ciency in nitrification processes (Bick 1972; Madoni, Davoli, and lyzed using the MetaMorph Imaging System software (Universal
Chierici 1993; Poole 1984). However, our group showed recently Imaging Corporation).
that several prostomatid species could be stable components in the Physical–chemical parameters were measured according to
biological communities of activated sludge plants with advanced standard methods (APHA 1989).
nitrogen removal systems (Pérez-Uz et al. 2009).
The aim of this work was to characterize a sessile prostomatid
ciliate found as a stable population associated with flocs in the
biological reactor of a WWTP from a health resort with a very RESULTS
specific set of physical–chemical conditions. We conclude that Specimens of the sessile prostomatid were found generally at-
this is a new species Metacystis galiani n. sp. tached on the surface or hidden in the activated sludge flocs (Fig. 1)
and are characterized below.
General morphology. Metacystis sp. is a flexible ciliate, ovoid
to bottle-shaped, covered by a corrugated sac-like lorica tapering
toward the anterior aperture. The cell morphology varies from
Corresponding Author: L. Arregui, Departamento de Microbiologı́a an elongated cell when the anterior oral area is extended (Fig. 2),
III, Facultad de Biologı́a, C/Jose Antonio Novais 2, 28040 Madrid, to an ovoid cell when this anterior part is retracted within the
Spain—Telephone number: 34 913 944 966; FAX number: 34 lorica (Fig. 3). Cell dimensions vary from 60.4 15.8 mm in
913 944 964; email: arregui@[Link] relaxed cells to 44.4 15.6 mm when contracted (Table 1). The
362
ARREGUI ET AL.—METACYSTIS GALIANI N. SP. 363

Fig. 1–6. General morphology of Metacystis galiani n. sp. 1. Metacystis galiani n. sp. embedded in activated sludge flocs. Silver carbonate staining,
scale bar 5 10 mm. 2. Living specimen inside the lorica. Note the extensible proboscide with the apical cytostome and the annulated lorica attached to the
flocs (phase contrast microscopy, scale bar 5 20 mm). 3. Phase contrast microphotography of a contracted cell, the protruding posterior vacuole is clearly
visible, scale bar 5 20 mm. 4, 5. General arrangement of the somatic and oral infraciliature of silver carbonate-stained cells, scale bars 5 20 mm.
6. Contractile vacuole pore (arrow) revealed with Flutax, scale bar 5 20 mm. Cy, cytostome; KF, kinetodesmal fiber; L, lorica; LR, lorica ridges; OC, oral
ciliature; P, proboscide; SK, somatic kineties; V, vacuole.
364 J. EUKARYOT. MICROBIOL., 57, NO. 4, JULY–AUGUST 2010

Table 1. Morphometric characterization of Metacystis galiani n. sp. Binary fission has been observed within the lorica, giving rise
to two ovoid-spherical cells, the anterior one slightly smaller than
Variable n Mean Standard Maximum Minimum the posterior one (Fig. 11, 12). The new opisthe oral structures
deviation originate by a telokinetal stomatogenesis (Fig. 12).
Relaxed cella
Ecology. Metacystis sp. was observed in the biological reactor
Length 14 60.4 5.1 67.6 51.6 of a WWTP from a freshwater health resort, where the water
Width 14 15.8 1.5 18.8 13.6 showed high conductivities and unusually high temperatures. The
Oral area 14 8.3 1.0 10.2 6.9 mixed water of the influent from the health resort is strongly min-
Contracted cella eralized with sulfates, bicarbonates, magnesium, and calcium as
Length 19 44.4 10.0 60.0 22.7 major ions and a minor content of lithium, manganese, zinc, lead,
Width 19 15.6 1.7 17.6 12.0 and chromium. Pre-treatment procedures also include Bayer cat-
Oral area 16 7.8 0.9 8.7 5.9 ionic exchange columns. Although the water originally flows from
Loricaa
a fountain at 14 1C, it is heated before it is employed and this is
Length 35 66.4 6.3 75.0 48.5
Width 35 20.3 1.8 24.4 16.1
why temperatures in the biological tank of the WWTP can be
Aperture 35 10.4 0.8 13.2 9.2 around 26–27 1C. The plant was characterized by mean organic
Ridges 36 13 1.4 16 11 load values of 274 mg O2/L of BOD5 (biological oxygen demand)
Number of longitudinal somatic kinetiesb and 556 mg O2/L of COD (chemical oxygen demand) and high
11 24.7 2.2 28 20 conductivity (2,469 mS/cm) due to the high values of SO24 ,
Number of transverse ciliary ringsb CO3H , Cl2 , Mg21, Ca21, and Na1 ions (Table 2).
19 7.8 0.6 9 7
a
Living specimens (measurements in mm). DISCUSSION
b
Stained cells.
This ciliate found in a sewage treatment plant has been assigned
to the genus Metacystis Cohn, 1866 because of its cell shape with
cytostome is located at the apical end of a retractile proboscide characteristic transverse ridges, the longitudinal arrangement of
(Fig. 2). One large vacuole protrudes at the terminal end of the cell somatic kineties, the apical circular cytostome with one circu-
(Fig. 2, 3, 13, 14). moral kinety and two rings of perioral cilia, the presence of a
Cilia pattern. Cells are uniformly ciliated by longitudinal posterior protruding vacuole, and the existence of a lorica attached
kineties in which the regularly disposed kinetosomes can be ob- to the substrate (Bick 1972; Carey 1992; Foissner 1984). The ge-
served as differentiated transverse rings of cilia (Fig. 4, 5, 14). nus Vasicola, certainly similar and within the same family, does
These cilia appear to maintain the cell on the ridges of the lorica, not present the protruding terminal vacuole as main differential
allowing it to move up and down within the lorica. No caudal character (Dragesco and Dragesco-Kernéis 1986).
cilium is present. Ciliates of the genus Metacystis are difficult to spot because of
Ciliature and infraciliature. The somatic infraciliature is their life style living inside a lorica, which is usually disguised
composed of 20–30 longitudinal kineties regularly disposed within sediment and other particles. This might be one of the rea-
around the body. Kineties are constituted of eight monokinetids, sons why some of these species have been poorly characterized.
uniformly spaced, giving rise to eight regular transverse girdles Three of them have been described in more detail employing
that constitute each of the paratenes (Fig. 4, 5). Each kinetid staining and electron microscopy methods: Metacystis striata
shows a thick kinetodesmal fiber (visualized in silver-impregnated (Foissner 1984), Metacystis truncata, and Metacystis borrori
specimens, Fig. 4, 5) and long postciliary and transverse micro- (Aladro-Lubel and Martı́nez-Murillo 1999, 2003). The character-
tubular fibers (detected with Flutax staining Fig. 6). A contractile istics observed in the species studied here, however, do not fit
vacuole pore opens between the fifth and sixth kinetids of one of within any of the species described in the genus Metacystis (Table
the somatic kineties (Fig. 6). 3). Differences with the other previously described species are
The circumoral infraciliature is composed of one circumoral related to the lorica shape, dimensions, cell infraciliature, pres-
kinety of dikinetids and two perioral girdles composed of the an- ence of a posterior protruding vacuole, and habitat.
terior ends of the somatic kineties. The anterior perioral fragment
is composed of two paratenes densely packed together while the (1) The sac-like lorica in this new Metacystis species is an-
more posterior perioral fragment is composed of four paratenes nulated with a lower number of girdles (11–16) than those
also densely packed (Fig. 7, 8, 14). observed in M. truncata (15–26). It also differs with this
Nuclear apparatus. Silver-impregnated cells show a single one in being smaller (Table 1).
macro- and micronucleus commonly located in the central body (2) The most similar species in relation to cell dimensions is
region. The macronucleus is almost spherical in shape, and the Metacystis elongata; however, Metacystis sp. has a lower
ellipsoid micronucleus is located in a small groove just beside the number of transverse ciliary rings (8 for the new species
macronucleus (Fig. 9, 14). against 18–21 for M. elongata) and one circumoral kinety
Structure of the lorica. The lorica of Metacystis sp. is a hy- of dikinetids and two perioral girdles, while M. elongata
aline structure usually found attached to the floc surface. Two has just one single perioral girdle (Table 3). Besides, a lo-
parts can be differentiated: a smooth anterior neck and an annula- rica has not been described for M. elongata.
ted body. The neck might be slightly curved and ends in a circular (3) While the 20–28 longitudinal somatic kineties found in
aperture 10.4 mm in diameter (Table 2). Mean values for lorica Metacystis sp. are similar to M. borrori, M. elongata, and
dimensions were 66.4 20.3 mm. The corrugated lorica body has Metacystis tesselata, the number of transverse ciliary rings
11–16 regularly spaced ridges and has a rounded bottom (Fig. 2, 3, differs: 8 vs. 5–7, 18–21, and 9 respectively. M. striata
10, 13). shows a considerably higher number of somatic kineties
Rounded particles, probably from digestive vacuoles, can often (i.e. 63–66). Moreover, the oral ciliature, both circumoral
be observed inside the lorica (Fig. 10). These particles sometimes and perioral, of Metacystis sp. is located in a characteristic,
are observed at the posterior pole while, in other cases, they are protrusible proboscide not mentioned by other authors for
laterally located, probably as cell activity pushed them out. other Metacystis spp.
ARREGUI ET AL.—METACYSTIS GALIANI N. SP. 365

Fig. 7–12. Detailed morphological aspects of Metacystis galiani n. sp. Scale bars 5 20 mm. 7, 8. Detail of the oral ciliature arrangement in a silver
carbonate (7) and Flutax (8) stained cells. Long postciliary fibers associated to the somatic kinetosomes revealed with Flutax staining. 9. Silver carbonate
stained cell showing the position of the macronucleus and micronucleus. 10. Rounded particles inside the lorica, probably egesta derived from food
vacuoles. 11. Phase contrast photomicrography of two daughter cells inside the lorica after binary fission. 12. Ciliature of two daughter cells stained with
Flutax showing telokinetal stomatogenesis. KF, kinetodesmal fiber; LN, lorica neck; LR, lorica ridges; Ma, macronucleus; Mi, micronucleus; OI, oral
infraciliature; PF, postciliary fiber; SP, secreted particles.
366 J. EUKARYOT. MICROBIOL., 57, NO. 4, JULY–AUGUST 2010

T, temperature; SV-30, sludge volume in 30 min; MLSS, mixed liquor suspended solids; SVI, sludge volumetric index; SS, suspended solids; BOD5, 5 days biological oxygen demand;
Total-P
(mg/L)

1.8
8.3

8.8
11.8
6.2
12.8
6.2
8.8
11.2
5.3
16.1
8
N-NH4
(mg/L)

52.8

12.9
33.7
12
25
50

50
46
14
30
56
22
Total-N
(mg/L)

65.7
56.9
50.4

36.2
66.1
31.4
19.8
40.3
16
30
52

19
Monthly influent average values

(mgO2/L)

555.8
COD

277
190
610
501
711
589
689
502
464
913
700
523
(mgO2/L)
BOD5

67
79
270
231
315
300
397
239
213
485
400
290
274
Fig. 13, 14. Schematic drawings of Metacystis galiani n. sp. 13. Mor-
phology of living cells. 14. Infraciliature. CI, circumoral infraciliature; Cy,
cytostome; KF, kinetodesmal fiber; LA, lorica aperture; LN, lorica neck;
LR, lorica ridges; Ma, macronucleus; Mi, micronucleus; OC, oral cilia-

COD, chemical oxygen demand; Total-N, total nitrogen; N-NH4, ammonium nitrogen; and Total-P, total phosphorous.
ture; P, proboscide; PF, postciliary fiber; PI1, perioral infraciliature 1; PI2,

(mg/L)

83
137

176
215
292
289
277
159
213
486
216
163
226
SS
perioral infraciliature 2; SK, somatic kineties; SP, secreted particles; V,
vacuole.

Conductivity
(mS/cm)
570
2,500
4,960
1,100
4,600
2,300
3,100
3,100
3,100
1,400
1,400
1,500
2,469
(4) The species studied has been isolated from samples of a
WWTP where it usually constituted the dominant popula-
Table 2. Mean values of physical–chemical parameters measured in the WWTP during 1 year.

tion. Species of this genus have been described from different

habitats—freshwater, marine, and brackish (Table 3)—but
they have never been reported as components of the protist
(u.d.)

community in a biological reactor of a WWTP. This makes

8.1
7.9
7.6
7.5
7.8
7.7
7.8
7.7
8.4
8.4
8.0
pH

8
8

the habitat of Metacystis sp. unique.

(ml/g)

Finally, several efforts have been devoted to amplify through

SVI

111
132
126
188
189
200
188
73
87
87
229
167
148

The establishment was opened to the public from March to November.

PCR, the ss-rRNA gene sequence of this Metacystis species. Since
no monoxenic cultures were successful, the process was tested
isolating ciliates from an original sample, and also with several
903
2,881
6,695
6,982
7,541
8,775

6,528
10,866
9,658
7,572
7,234
5,859
3,368
(m3/d)

specimens fixed with Bouin. Unfortunately, no results have been

Flow

obtained until now but this work is ongoing and it is expected that
the sequence will be submitted to GenBank as soon as the ampli-
fication procedure is successful.
Aeration tank

In conclusion, we propose this species as a new species named

(mg/L)
MLSS

4,954
4,533
3,953
4,782
5,300
3,500
3,715
4,404
3,460
3,446
3,493
4,800
4,195

M. galiani n. sp.
Class Prostomatea
Order Prostomatida
SV-30

550
600
500
900

700
700
320
300
300
800
800
623
1,000

Family Metacystidae
(ml)

Metacystis galiani [Link]. (Table 1, Fig. 1–14)

Diagnosis. In vivo ovoid cells 22.7–60.0 mm (mean value

44.4 mm) 12.0–17.6 mm (mean value 15.6 mm) to bottle shaped
T (1C)

26.3

cells 51.6–67.6 mm (mean value 60.4 mm) 13.6–18.8 (mean

12
23
23
29
30
30
30
30
29
25
30
24

value 15.8 mm). Anterior cytostome located at the apical end of

a retractile proboscide. Cells placed in a corrugated sac-like lorica
48.5–75.0 mm (mean value 66.4 mm) 16.1–24.4 mm (mean value
Mean values

20.3 mm), a hyaline structure with a smooth anterior neck and an

September

November
December
February

annulated body (11–16 ridges). Somatic ciliature formed by lon-

October
January

August
March
April

gitudinal somatic kineties (20–30), each constituted by eight

May
June
July

kinetids in paratenes. Oral infraciliature with one circumoral

Table 3. Comparative data of different Metacystis species.

Species Description Shape Size (mm) Cytostome Number of Number of Circumoral Ma Mi Terminal Caudal Lorica Habitat
somatic transverse ciliature vacuole cilium
Shape Size (mm) Number
kineties ciliary
of
rings
girdles
Metacystis Stokes, 1895 Elongate 55 _ _ _ _ _ _ Present Absent Cylindrical, 140 Present Brackish
annulata with
anterior
neck
Metacystis Aladro-Lubel Ovoid- 10–35 10–18 Central 22–30 5–7 2 Perioral 3–7 mm Absent Cylindrical 18–61 Absent Marine
borrori & Martı́nez- cylindrical 1 circumoral 11–26
Murillo, 2003
Metacystis Penard, 1922 Elongate 75–170 _ _ 24 _ _ _ Present Absent Tubular, _ _ Freshwater
daphnicola curved
anteriorly
Metacystis Kahl, 1928 Cylindrical, 40–120 10 Central 30 18–21 1 Perioral 5.5 mm 1 Present Absent _ _ _ Marine,
elongata elongate brackish
Metacystis Penard, 1922 Saccate 13–20 Acentric _ 5–6 2 Perioral _ _ Absent Various Saccate _ Absent Freshwater
exigua
Metacystis Elongate to 44–60.5 15–16 Apical 20–28 8 2 (412) Single, Single, Present Absent Sac-like 66.4 20 11–16 Biological
galiani [Link]. ovoid Perioral ovoid ellipsoid reactor
1 circumoral of
(dikinetids) WWTP
Meacystis Dietz, 1964 Elongate 92 _ _ 4 _ _ _ Present Absent Like M. _ _ Marine,
hyperhalina annulata
Metacystis Penard, 1922 Vase-like 30 Acentric Present _ 2 Perioral Small 4 Absent Various Vase-like 50–62 Absent Freshwater
lagenula
Metacystis Kahl, 1930 Elongate 80–120 _ _ _ _ _ _ Absent Absent Cylindrical, _ _ Freshwater
megapterae semiannular,
rounded
ARREGUI ET AL.—METACYSTIS GALIANI N. SP.

posteriorly
Metacystis Penard, 1922 Elongate 45–68 15 1 Present 1 Vase-like Freshwater,
recurva marine
Metacystis Stokes, 1893 Cylindrical 50–110 25–39 Central 63–66 20–24 2 Perioral 1 10 mm 1 Present Absent Tubular 100 48 Absent Brackish,
striata circumoral marine
Metacystis Kahl, 1926 Cylindrical 50 Central 20–26 9 1 Perioral 1 Present 1 Vase-like 84 25 Absent Freshwater
tesselata
Metacystis Cohn, 1866 Ovoid- 10–91 11–30 Central 10–18 Abundant 7– 1 Present Absent Cylindrical, 77–140 15–26 Brackish,
truncata cylindrical perioral 14 mm annular 24.5–31.5 marine
cilia

Data obtained from Aladro-Lubel and Martı́nez-Murillo (1999, 2003), Borror (1972); Carey (1992); Dragesco and Dragesco-Kernéis (1986); Foissner (1984); Kahl (1930).
367
368 J. EUKARYOT. MICROBIOL., 57, NO. 4, JULY–AUGUST 2010

kinety and two perioral girdles; each perioral fragment constituted Curds, C. R. 1969. An illustrated key to the British freshwater ciliated
by two and four densely packed paratenes, respectively. One large protozoa commonly found in activated sludge. Wat. Pollut. Res., Tech-
vacuole protruding at the posterior end with its pore located be- nical paper, 12:1–90.
tween the fifth and sixth kinetids of one of the somatic kineties. Curds, C. R. & Cockburn, A. 1970. Protozoa in biological sewage treat-
ment processes. II. Protozoa as indicators in the activated sludge pro-
Dedication. We dedicate this new species to the memory of
cess. Water Res., 4:237–249.
Dr. Dimas Fernández-Galiano, a distinguished Spanish proto- Dragesco, J. & Dragesco-Kernéis, A. 1986. Ciliés libres de l’Afrique
zoologist and our professor for many years. intertropicale. Editions de l’Orstom, Paris.
Type locality. Adhered on the surface or camouflaged into the Esteban, G., Téllez, C. & Bautista, L. M. 1991. Dynamics of ciliated pro-
activated sludge flocs from an aerobic biological reactor of a tozoa communities in activated sludge process. Water Res., 25:967–
WWTP from a health resort located in Cofrentes, Valencia, Spain 972.
(39113 0 N, 1104 0 W). Fernández-Galiano, D. 1976. Silver impregnation of ciliated protozoa:
Type material. Permanent silver-stained preparations were procedure yielding good results with the pyridinated silver carbonate
deposited at the Museo Nacional de Ciencias Naturales (MNCN, method. Trans. Am. Micros. Soc., 95:557–560.
Madrid, Spain). Accession number: MNCN 39.04/1. Fernández-Galiano, D. 1994. The ammoniacal silver carbonate method as
a general procedure in the study of protozoa of sewage (and other)
waters. Water Res., 28:495–496.
ACKNOWLEDGMENTS Foissner, W. 1984. Infraciliatur, Silberliniensystem und Biometrie einiger
neurer und wenig bekannter terrestrischer, limnischer und mariner Cil-
The authors would like to thank Da Dolores Fernández Abellán iaten (Protozoa, Ciliophora) aus den Klassen Kinetofragminophora,
(Egevasa), who is responsible for the operation of the wastewater Colpodea und Polyhymenophora. Stapfia, Linz, 12:1–165.
treatment plant for her interest in this study and her contribution of Kahl, A. 1930. Urtiere oder Protozoa I: Wimpertiere oder Ciliata (In-
data. We also acknowledge D. Rafael Lloret Salinas (EPSAR, fusoria). Allgemeiner Teil und Prostomata. In: Dalh, F. (ed.), Die Tie-
Entidad Pública de Saneamiento de la Generalidad Valenciana) rwelt Deutschlands und der angrenzenden Meeresteile. Gustav Fischer,
and D. Vicente Fajardo Montañana (EGEVASA) for their atten- Jena. p. 1–180.
tion, the physical–chemical parameters provided, and the contin- Laspidou, C. S. & Rittmann, B. E. 2002. A unified theory for extracellular
ued support of these companies to research on activated sludge polymeric substances, soluble microbial products, and active and inert
biomass. Water Res., 36:2711–2720.
processes. This work was supported by grant CGL2008-02310 Madoni, P. 1988. I protozoi ciliate nel controllo di efficienza dei
from the Spanish Ministerio de Ciencia e Innovación. fanghi attivi. Centro Italiano Studi di Biologia Ambientale, Reggio
Emilia.
LITERATURE CITED Madoni, P., Davoli, D. & Chierici, E. 1993. Comparative analysis of the
activated sludge microfauna in several sewage treatment works. Water
Aladro-Lubel, M. A. & Martı́nez-Murillo, M. E. 1999. First description of Res., 27:1485–1491.
the lorica of Metacystis truncata and its occurrence on Thalassia Martı́n-Cereceda, M., Serrano, S. & Guinea, A. 1996. A comparative study
testudinum. J. Eukaryot. Microbiol., 46:311–317. of ciliated protozoa communities in activated sludge plants. FEMS
Aladro-Lubel, M. A. & Martı́nez-Murillo, M. E. 2003. Metacystis borrori Microbiol. Ecol., 21:267–276.
n. sp. (Ciliophora: Metacystidae) on the Seagrass Thalassia testudinum. Martı́n-Cereceda, M., Serrano, S. & Guinea, A. 2001. Biofilm communi-
J. Eukaryot. Microbiol., 50:204–209. ties and operational monitoring of a rotating biological contactor sys-
APHA. 1989. Standard Methods for the Examination of Water and Waste- tem. Water Air Soil Poll., 126:193–206.
water. 17th ed. American Public Health Assocciation, Washington, DC. Pérez-Uz, B., Arregui, L., Calvo, P., Salvadó, H., Fernández, N., Rodri-
Arregui, L., Muñoz, C., Guinea, A. & Serrano, S. 2003. FLUTAX em- guez, E., Zornoza, A. & Serrano, S. 2009. Efficiency of nitrogen re-
ployment simplifies the visualization of the ciliature of oxytrichid hypo- moval and protist communities: the potential for introduction of novel
trichs. Eur. J. Protistol., 39:169–172. biological index. Proceedings of the International Workshop: Integrated
Barker, D. J. & Stuckey, D. C. 1999. A review of soluble microbial prod- vision of urban and agro-industrial wastewater treatment, monitoring
ucts (SMP) in waste water treatment systems. Water Res., 33:3063– and reclamation: the key role played by the sewage treatment plant,
3082. Terni, Italy.
Bick, H. 1972. Ciliated Protozoa. An Illustrated Guide to the Species used Poole, S. E. P. 1984. A study of the relationship between the mixed liquor
as Biological Indicators in Freshwater Biology. World Health Organi- fauna and plant performance for a variety of activated sludge sewage
zation, Geneva. treatment works. Water Res., 18:281–287.
Bitton, G. 2005. Wastewater Microbiology. 3rd ed. Wiley-Liss Publisher, Wilbert, N. 1975. Eine verbesserte Technik der Protargolimprägnation für
New York. Ciliaten. Mikrokosmos, 64:171–179.
Borror, A. C. 1972. Tidal marsh ciliates (Protozoa): morphology, ecology,
systematics. Acta Protozool., 10:29–71.
Carey, P. G. 1992. Marine Insterstitial Ciliates. An Illustrated Key. Chap-
man & Hall, London. Received: 11/16/09, 02/04/10, 03/18/10; accepted: 03/20/10