Unusual Features of Non-Dividing Somatic Macronuclei in the Ciliate Class Karyorelictea

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European Journal of Protistology 61 (2017) 399–408

REVIEW

Unusual features of non-dividing somatic macronuclei in the ciliate class Karyorelictea

a,b b a b,c,∗

Ying Yan , Anna J. Rogers , Feng Gao , Laura A. Katz

Laboratory of Protozoology, Institute of Evolution and Marine Biodiversity, Ocean University of China, Qingdao 266003, China

Department of Biological Sciences, Smith College, Northampton, MA 01063, USA

Program in Organismic and Evolutionary Biology, University of Massachusetts, Amherst, MA 01003, USA

Available online 22 May 2017

Abstract

Genome structure and nuclear organization have been intensely studied in model ciliates such as Tetrahymena and Paramecium,

yet few studies have focused on nuclear features of other ciliate clades including the class Karyorelictea. In most ciliates, both

the somatic macronuclei and germline micronuclei divide during cell division and macronuclear development only occurs after

conjugation. However, the macronuclei of Karyorelictea are non-dividing (i.e. division minus (Div−)) and develop anew from

micronuclei during each asexual division. As macronuclei age within Karyorelictea, they undergo changes in morphology and

DNA content until they are eventually degraded and replaced by newly developed macronuclei. No less than two macronuclei

and one micronucleus are present in karyorelictid species, which suggests that a mature macronucleus 1) might be needed

to sustain the cell while a new macronucleus is developing and 2) likely plays a role in guiding the development of the new

macronucleus. Here we use a phylogenetic framework to compile information on the morphology and development of nuclei in

Karyorelictea, largely relying on the work of Dr. Igor Raikov (1932–1998). We synthesize data to speculate on the functional

implications of key features of Karyorelictea including the presence of at least two macronuclei in each cell and the inability

for macronuclei to divide.

creativecommons.org/licenses/by-nc-nd/4.0/).

Keywords: Karyorelictea; Non-dividing macronuclei; Nuclear architecture

Introduction and Blackburn 1985). Yet the bulk of the work on ciliates

has predominately focused on only a few lineages, such as

Ciliates are a clade of single-celled eukaryotes deﬁned by Paramecium and Tetrahymena (Class Oligohymenophorea;

the presence of cilia and dimorphic nuclei (Kovaleva and Aury et al. 2006; Chalker 2008; Eisen et al. 2006; Mochizuki

Raikov 1992; Prescott 1994; Raikov 1982, 1985). Numerous and Gorovsky 2004; Yao 2008) and Oxytricha and Stylony-

key discoveries have been made through studies of cili- chia (Class Spirotrichea; Chen et al. 2014; Nowacki et al.

ates, including the discovery of self-splicing RNA (Cech 2008).

1986; Kruger et al. 1982) and telomeres/telomerases (Greider As described in detail below, the monophyletic class

Karyorelictea, the focus of this manuscript, appears unique

∗ among ciliates in that their somatic macronuclei cannot divide

Corresponding author at: Department of Biological Sciences, Smith Col-

(Table 1; Raikov 1982). Instead, Karyorelictea differentiate

lege, 44 College Lane, Northampton, MA 01063, USA. Fax: +1 413 585

3786. new macronuclei from micronuclei with each cell division

E-mail address: [email protected] (L.A. Katz).

http://dx.doi.org/10.1016/j.ejop.2017.05.002

400 Y. Yan et al. / European Journal of Protistology 61 (2017) 399–408

Table 1. Unusual nuclear features in karyorelictids in comparisons with other major ciliates classes.

a c

Subphylum Class Mac Div Mac Mac differentation Mac aging b

differentation during asexual

post conjugation division

Postciliodesmatophora Karyorelictea Div− + + +

Heterotrichea Div+ + − −

Intramacronucleata Litostomatea Div+ + − −

− −

Armophorea Div+ +

Spirotrichea Div+ + − −

Colpodea Div+ + − −

Nassophorea Div+ + − −

Phyllopharyngea Div+ + − −

Prostomatea Div+ + − −

Plagiopylea Div+ + − −

Oligohymenophorea Div+ + − −

Ability of macronuclear division. b Macronuclear differentiation.

Macronuclei age as they persist through several generations without dividing (see text for more detail).

and an individual macronucleus only persists for a few gen- tivable, the genus Loxodes has been cultivated successfully

erations before degrading (Table 1; Raikov 1985). In all other in a few labs (e.g. Bobyleva 1981; Buonanno et al. 2005;

ciliates, the macronuclei divide by amitosis (i.e. dividing Finlay et al. 1986), likely accounting for the limited data on

without centromeres and metaphase plates) during vegetative this clade.

growth and only differentiate from zygotic nuclei following Here we describe some unusual features within the class

conjugation (e.g. Gao et al. 2016; Raikov 1982). Through- Karyorelictea, focusing on their somatic macronuclei. We

out the remainder of this manuscript, we use the abbreviation divide the manuscript into three parts: 1) a description of the

−

Div to describe the absence of macronuclear division in contributions from Dr. Igor Raikov, who generated the bulk

Karyorelictea; all remaining classes of ciliates are Div+ of data on members of the class Karyorelictea; 2) insights into

as they contain macronuclei that divide through amitosis the cell biology of the Karyorelictea based on microscopic

(Table 1). analyses; and 3) knowledge of life cycles, and particularly

The class Karyorelictea is composed of ∼170 ciliate macronuclear development, in diverse lineages within the

morphospecies divided into six families (Loxodidae, Trach- class Karyorelictea. Finally, we end with hypotheses on the

elocercidae, Kentrophoridae, Geleiidae, Cryptopharyngidae evolutionary signiﬁcance of Div− macronuclei.

and Wilbertomorphidae; Fig. 1; Lynn 2008). However,

limited molecular data are available in karyorelictids; for

Raikov and Karyorelictea

example, as we write this manuscript, only eight protein-

coding genes have been released on GenBank for the

Dr. Igor B. Raikov (1932–1998) was the leading researcher

class Karyorelictea. The current phylogeny of Karyorelictea

in the study of Karyorelictea, and most information about

has largely relied on small subunit rDNA sequences (e.g.

these ciliates stems from his work and the work of his col-

Andreoli et al. 2009; Campello-Nunes et al. 2015; Gao et al.

laborators. Raikov characterized in detail the ﬁne structure

2016; Xu et al. 2015; Yan et al. 2016), with the excep-

of nuclei for more than 20 karyorelictid species, including

tion of the family Cryptopharyngidae for which there are

representatives from four of the six families in Karyorelictea

currently no molecular data. Phylogenetic analyses indicate

(e.g. Kovaleva and Raikov 1992; Raikov 1982, 1985; Raikov

that both the class Karyorelictea and its ﬁve families with

and Karadzhan 1985; Raikov and Kovaleva 1990). Using an

molecular data are monophyletic and fall sister to the class

impulse cytoﬂuorimeter, Raikov isolated individual nuclei

Heterotrichea, the other member of the subphylum Postcil-

and measured their DNA content. From these data on the

iodesmatophora (Campello-Nunes et al. 2015; Gao and Katz

morphology and DNA content of karyorelictid nuclei, Raikov

2014; Gao et al. 2016; Xu et al. 2015; Yan et al. 2016). Most

and his collaborator, Dr. Valentine G. Kovaleva, described the

karyorelictids are specialized to interstitial marine environ- −

Div macronuclei as “paradiploid”, implying that mature

ments such as sand, and recently they were also found to be

macronuclear are nearly diploid and similar to micronuclei

epibionts of benthic harpacticoid copepods in the deep sea

in DNA content (Kovaleva and Raikov 1978; Ovchinnikova

(Sedlacek et al. 2013); the one exception is the genus Lox-

et al. 1965), which is unusual as the macronuclei in other

odes, which is found in low oxygen freshwater environments

ciliate species can be highly polyploid (for example 45 in

(Fenchel 1967, 1968; Finlay and Fenchel 1986). Additionally,

Tetrahymena, 800 in Paramecium and ∼13150 in Spirosto-

while none of the rest of Karyorelictea are reported as cul-

mum; Duret et al. 2008; Ovchinnikova et al. 1965; Woodard

Y. Yan et al. / European Journal of Protistology 61 (2017) 399–408 401

Fig. 1. A cartoon tree of Karyorelictea showing main diversity of nuclear clustering within and among families based on small subunit rDNA

(SSU-rDNA) maximum likelihood tree. Cell sizes are drawn roughly to scale. There are currently no molecular data on Cryptopharyngidae

(*); thus it is displayed as an orphan family. Scale bar: 100 ␮m.

et al. 1972). Raikov’s many other contributions are high- be nuclei enclosed by a fused membrane or nuclei that

lighted in some of the work we review below. are closely associated with one another. A unit of two

macronuclei and one micronucleus is the smallest and most

frequent grouping, though groups of four macronuclei and

two micronuclei are also commonly described in karyore-

Part I: nuclear structure

lictid species (Raikov 1994). Many karyorelictids possess

multiple groups of either 2 + 1 or 4 + 2 units (e.g. Trachelo-

Insights from light microscopy

raphis oligostriata and T. colubis; Fig. 1; Al-Rasheid 2001;

As in all other ciliates, karyorelictids have both somatic

Xu et al. 2011b). Free nuclei, which are common in other

macronuclei and germline micronuclei in every individ-

multinucleate ciliates, are also often found in multinucle-

ual cell. The majority of macronuclei and micronuclei in

ate karyorelictids (e.g. Kentrophoros ﬂavus and K. gracilis;

Karyorelictea are spherical or elliptical, and are usually

Fig. 1), and in many cases, these free nuclei form a longitu-

5–10 ␮m (macronuclei) and 2–4 ␮m (micronuclei) in diam-

dinal line along the cell meridian (Fig. 1; Xu et al. 2011a).

eter or length (Raikov 1985). Karyorelictean ciliates with

In karyorelictids, nuclear grouping and number of nuclei

only one macronucleus have not yet been found, while this

may relate to body shape and size in some, but not all lineages

arrangement of nuclei is common in all other ciliate classes

(Table 2). In Loxodes and Remanella (Loxodidae), larger

(Lynn 2008; Raikov 1982). This suggests that each karyore-

cells tend to have more macronuclei and micronuclei, but

lictean cell requires a minimum of two macronuclei and one

the same pattern is not found in Trachelocercidae (Table 2).

micronucleus to survive (Andreoli et al. 2009; Lynn 2008;

For example, both Trachelocerca sagitta and Trachelocerca

Raikov 1982).

stephani have single nuclear groups with 4 macronuclei and

Nuclei in karyorelictids can be distributed throughout

2 micronuclei; however, the size of T. sagitta is ∼5 fold that

the cell body or clustered together as “nuclear groups” or

of T. stephani (Table 2). There is also considerable variation

“nuclear complexes” (Fig. 1). Nuclear groups can either

402 Y. Yan et al. / European Journal of Protistology 61 (2017) 399–408 (1996) (1996) (1996) (1996) (1989)

(1983) (1983)

Dragesco Dragesco Dragesco Dragesco Rieder Rieder

Volkonitin (2001)

(2002) (2002) (2002) (2016) (2016) (2013)

(2011b) (2013) (1995) and and and and (1996c) (1996c) and and (1996b) (1996a)

(1994) and

(1973)

al. al. al.

al. al.

et et et

et et

Foissner Foissner Epstein Foissner Raikov Yan Yan Borror Al-Rasheid Dragesco Foissner Foissner Dragesco Dragesco Xu Yan Foissner Foissner Xu Foissner Foissner Foissner Foissner References

mic

group mic

11–54 6–12 5–37

mic mic

× × ×

mic

mic mic mic

3–9 5–12 3–7 10–16

mic mic mic mic 2 2 2

+ + + + mic mic

mic) mic) mic) 1 2

1 1–3

+ + + mic mic mic mic mic mic

3 6

2 1 2 + + + +

2–3 2 2 2 2 1 2 micronucleus.

+ +

+ + + mac mac mac mac

+ + + + + + +

mac mac mac

nuclei/nuclear mac mac mac mac

mac mac Mic, mac mac mac

mac mac mac mac mac of mac mac

6–12 (4 3 7–14 2–3 4 4 4 4 10 2–3 17–39 (4 10–24 3–5 2 2 18–26 16 # m ␮

a macronucleus.

3 2 5 4.5

× × × ×

a 2 6 3.6 3 6 2.2 4 2.6–2.9 2.8–3.6 5 2 3–5 1.5–2 Mic diameter/ Mac,

m ␮ species.

5.5 6.3 2.2

3.5 52 52 4 6

× ×

10 2–2.6 5–12 × × × × ×

4 7–105–8 2–4 2–3 3–4 (2 ∼ 7–10 3–6 7.9 8 6 12.5 8 7.3 10 3–5 5–12 2-3 5–6 6.6–9.3 7–8 Mac diameter/

karyorelictid

m ␮

diverse

width/

20–30 30 35 62.7 87 90 30 – 50 41–50 80–84 40–80 10–20 25 15 30–40 Cell

group

m ␮

length/

nuclei/nuclear 500–2000 1100–2300 50–60 400–9001000–1500 40–60 30–50 8 6–8 4 285–500 62–75 5–6 2–3 15–43 1300 400–500 15 340 700 1500–2000 75–100 10 437 800–10001000 40–50 10 500–1000 800–1250 300–600 800–1500 1000–3000 180 Cell 209–250 340–440 45 50–90

number

colpoda

aragoi haloetes

obscura incaudata ditis ditis sagitta stephani and

ﬁstulosus

sulcata multinucleata

size

striatus magnus

cell name

ﬁgure.

Tracheloraphis Kentrophoros Tracheloraphis primitarum Tracheloraphis phoenicopterus Wilbertomorpha Trachelolophos quadrinucleatus Tracheloraphis Tracheloraphis oligostriata Trachelocerca Trachelocerca Trachelolophos binucleatus Trachelocerca Trachelocerca Trachelocerca Trachelocerca Apotrachelocerca arenicola Kovalevaia Prototrachelocerca caudata Prototrachelocerca fasciolata Loxodes Loxodes Remanella Taxa Apocryptopharynx hippocampoides table from

estimated

Summary is

2. available.

not

, Number Kentrophoridae Trachelocercidae Family Wilbertomorphidae Cryptopharyngidae Loxodidae − a Table

Y. Yan et al. / European Journal of Protistology 61 (2017) 399–408 403

Fig. 2. Examples of karyorelictid macronuclei showing different composition of nucleoli (1), crystalloid bodies (2), chromocenters (3) and

unknown spheres (4): A. macronucleus with a single nucleolus and crystalloid plus numerous chromatin bodies (arrowheads) from Trache-

loraphis caudatus (modiﬁed from Raikov 1994). B. macronucleus with a large chromocenter, nucleolus and sphere found in Trachelocerca

multinucleata (modiﬁed from Raikov 1989). C. macronucleus with a large number of small nucleoli in Geleia orbis (modiﬁed from Raikov

1984). D. simple macronucleus with multiple nucleoli Remanella granulosa (modiﬁed from SEM in Raikov and Kovaleva 1990).

within species. For example, the length of Prototrachelocerca and are also typically less electron dense than the ﬁbrillar

␮

fasciolata varies between 1000 and 3000 m with 10–24 core of nucleoli (Raikov 1990). No consistent association

macronuclei and 3–7 micronuclei, and Loxodes magnus can between nuclear bodies (spheres or crystalloids) and other

have 8–31 macronuclei and 5–32 micronuclei (Bobyleva et al. nuclear structures has been found in karyorelictean macronu-

1980). clei (Table 3; Raikov 1985, 1990).

Like in other ciliates, no nucleoli nor evidence of RNA syn-

thesis has been found in karyorelictean micronuclei during

Insights from electron microscopy

vegetative stages (Raikov 1982). Compared to macronu-

Fine structure analyses of karyorelictid nuclei reveal vari-

clei, micronuclei stain intensely and homogeneously with

ability in compositions of macronuclei while micronuclei

Feulgen indicating that they are packed with condensed chro-

have more canonical eukaryotic features (Raikov 1985,

matin (Raikov 1985). In Remanella multinucleata and Geleia

1994). In contrast to other ciliates, the somatic macronuclei of

orbis, small nuclear bodies that appear to be crystalloids

Karyorelictea are relatively small, ranging from 5 to 10 ␮m in

are sometimes found in the micronuclear periphery (i.e. the

diameter (Raikov 1982, 1985). Karyorelictean macronuclei

space between the chromatin mass and the nuclear envelope;

are also described as the “vesicular type” due to their shape,

Kovaleva and Raikov 1978; Raikov 1979, 1984).

and they often include nucleoli, chromocenters (i.e. areas of

condensed chromatin), and “nuclear bodies” (Raikov 1985,

1989). Nucleoli in karyorelictids have a similar ultrastructure Part II: development of macronuclei

to other eukaryotes and are described as having a classical ﬁb-

rillar core and granular cortex (Raikov 1985). Nucleoli serve Nuclear reorganization

as the location for the generation of rRNAs (Raikov 1982). The common ancestor of Karyorelictea evolved a

The number of nucleoli varies among karyorelictid species mechanism to segregate nuclei between generations by differ-

from one to over 100 (Fig. 2A–D; Raikov 1985). entiating at least one new macronucleus from a micronucleus

Chromocenters and chromatin bodies are two types of with each division (Orias 1991a). This contrasts with all other

densely staining regions of chromatin varying by size and ciliates in which macronuclei divide through amitosis either

number (Fig. 2A). Chromatin bodies are usually smaller and with microtubules placed inside or outside the macronuclear

more numerous than chromocenters, and chromocenters may envelope (subphylum Intramacronucleata and the class Het-

be formed by the fusion of chromatin bodies (Fig. 2A; Raikov erotrichea, respectively; Lynn 2008).

1979, 1990, 1994). Chromatin bodies may represent lowly Division patterns of Loxodes and lineages with two other

expressed heterochromatin regions, but the functional impli- common nuclear group types in karyorelictids exemplify the

cations of chromocenters are as yet unknown. diversity of processes within this class. In the simplest case

Another unusual aspect of the ﬁne structure of some kary- of two macronuclei and one micronucleus in each cell (e.g.

orelictean macronuclei is the presence of nuclear bodies, Loxodes rostrum; Fig. 3A–D), the two parental macronuclei

which are either classiﬁed as spheres or crystalloids (Fig. 2A, are separated into the daughter cells while the micronucleus

B; Table 3; Raikov 1979, 1984, 1985, 1990). These nuclear undergoes mitosis twice (Fig. 3A–C) and two micronuclei

bodies are of unknown function and are not present in all from the second mitosis differentiate into new macronu-

Karyorelictea: for example, they have not been described in clei (Fig. 3C, D; Raikov 1957). In species with two nuclear

the genus Loxodes (Table 3; Raikov 1979, 1990). Crystalloids groups, each with one macronucleus and one micronu-

are typically composed of some type of proteinaceous ﬁla- cleus (e.g. Loxodes striatus), each micronucleus divides once

ments, packed tightly together and parallel, and are usually (Fig. 3E, F), with one of the daughter micronuclei develop-

less electron dense than the ﬁbrillar core of nucleoli (Raikov ing into the new macronucleus while the other divides again

1979, 1985, 1990, 1994). In spheres, ﬁlaments are tangled to reset micronuclear number of the daughter cells (Fig. 3G,

404 Y. Yan et al. / European Journal of Protistology 61 (2017) 399–408

H; Raikov 1957). In species with four macronuclei and two

micronuclei in a nuclear group (e.g. Tracheloraphis colu- (1985)

(1978) (1980) (1996)

ber), each nuclear group is divided in half and then each half

(two macronuclei and one micronucleus) undergoes similar (1981)

developmental processes to form a complete nuclear group (1989)

al. Kovaleva Kovaleva Karadzhan Kovaleva

(four macronuclei and two micronuclei) after micronuclei al. et

(1985) (1979) and and and et and (1984) divide twice (Fig. 3I–L; Raikov 1969). Instead of generating

a completely new nuclear group in one of the daughter cells,

karyorelictean species that divide by the third method effec- References

Raikov Raikov Raikov Raikov Raikov Raikov Bobyleva Raikov Raikov

tively decrease the age difference of nuclei between daughter

cells, i.e. if a completely new nuclear group is generated and

passed on to one of the daughter cells while the other one

receives the parental nuclear group, there is a one-generation with

chromatin age difference between the offspring and this age difference

with aging/other

macronuclei,

matrix

would increase each division.

and

with chromocenters

chromocenters nucleoli, nucleoli with

aging contact with

Development of macronuclei continuous

in in

Given the inability of karyorelictean macronuclei to divide

−

(i.e they are Div ), nearly half of the macronuclei in each contact

occasionally bodies chromocenters structures nucleoli, N/A None continuous

matrix cell are newly differentiated from micronuclei following cell

division. As described below, the general pattern of macronu-

clear development in karyorelictean species appears similar

to other ciliates except that no evidence for genomic ampliﬁ-

nmnm Sometimes – nm Always cation during development has been recorded (Raikov 1957,

width Associations

observable

nmnm In nm Sometimes Sometimes

1959, 1982). Macronuclear development begins with cre- nm Always

Fiber N/A Not

7–8 7–8 10–12 10–12 7–8 ation of vacuoles in the developing macronucleus (Fig. 4A),

which is followed by the formation of chromatin granules

(Fig. 4B). In some species the size and number of chromatin m 10–12 ␮

granules decrease during development (e.g. Tracheloraphis 8

to margaritatus; Fig. 4C; Raikov 1957), suggesting that DNA

elimination may be occurring (Raikov 1994). DNA elimi-

nation is a common aspect of macronuclear development in

␮

species.

other ciliates where regions of germline DNA are selectively ␮

Diameter

N/A – –

– – 2

excised or deleted (Prescott 1994). Later, the emergence of

nucleoli indicates the maturation of macronuclei (Fig. 4D;

Raikov 1985). The few studies focusing on conjugation in

RNA RNA – RNA – RNA 2–3

karyorelictid

Karyorelictea show differences compared to dividing cells;

for instance, macronuclear development is blocked during

Material N/A Protein Protein Protein Protein Protein Protein – Protein conjugation and resumes only after cells separate (Raikov diverse

in 1990).

Aging of macronuclei

Since karyorelictean macronuclei cannot divide, they age, inclusions

and the macronuclei within a cell are always at different ages

Crystal/sphere Sphere Sphere Sphere Sphere Sphere Sphere

None Sphere (Raikov 1982). As macronuclei age, changes occur in their

size, shape, staining properties, and ﬁne structure (Raikov

1985). The size of young macronuclei is often between that macronuclear

of micronuclei and mature macronuclei; for example, in of

Trachelocerca multinucleata, micronuclei are measured at

table

␮ ␮

5–6 m in diameter, and young macronuclei are 5–7 m,

while mature macronuclei are 8–10 ␮m in diameter (Raikov phoenicopterus dogieli crassus

obscura geopetiti

tubiformis

1989). Young macronuclei also stain less strongly with Feul- multinucleata

Summary gen than mature or old macronuclei, and possess smaller

applicable.

orbis

available. and more scattered chromatin bodies (Bobyleva et al. 1980;

not

not Kovaleva and Raikov 1978; Raikov and Kovaleva 1996). In

, − Table TaxonLoxodes Inclusion

Remanella Tracheloraphis Tracheloraphis N/A,

Tracheloraphis Trachelocerca Trachelocerca Kentrophoros Geleia

contrast, mature macronuclei stain darker and have larger

Y. Yan et al. / European Journal of Protistology 61 (2017) 399–408 405

Fig. 3. Nuclear reorganization during cell division in karyorelictids. A–D. Pattern of nuclear group with two macronuclei and one micronucleus

(modiﬁed from Loxodes rostrum, Raikov 1957): two parental macronuclei are segregated into daughter cells and each daughter cell receives

a newly differentiated macronucleus. E–H. Pattern for a nuclear group with two macronuclei and two micronuclei (modiﬁed from L. striatus,

Raikov 1957): similar to the ﬁrst pattern with an additional micronuclear division. I–L. Pattern for nuclear group with four macronuclei and

two micronuclei (modiﬁed from Trachelocerca coluber, Raikov 1969): the parental nuclear group is separated into two halves, and the full

complement of nuclei is restored by nuclear division and differentiation.

Fig. 4. Macronuclear development in Tracheloraphis margaritatus (Feulgen staining, modiﬁed from Raikov 1957). A. vacuolization stage

from a macronuclear anlagen. B. macronuclear anlagen forming numerous chromatin granules. C. dechromatinization stage. D. appearance

of nucleoli.

chromocenters (Bobyleva et al. 1980; Kovaleva and Raikov et al. 1980; Kovaleva and Raikov 1978; Raikov et al. 1963).

1978; Raikov and Kovaleva 1996). Mature macronuclei tend The functional implications of changing DNA content are

to be relatively more abundant in multinuclear cells (Raikov unknown.

1985). Old macronuclei stain darker still and have more The Div− macronuclei in karyorelictids also have a

irregular shapes with a higher presence of spheres in some relatively short life span. They persist only through 3–7

species (e.g. Kentrophoros tubiformis and Kovalevaia (Trach- cell divisions as estimated from the percentage of degen-

elonema) sulcata; Bobyleva et al. 1980; Kovaleva and Raikov erating macronuclei per cell in Loxodes magnus, while

1978; Raikov and Kovaleva 1996). macronuclei appear to exist longer in Kentrophoros ﬁstulo-

DNA accumulation in macronuclei accompanies the mor- sum (Raikov 1985, 1994; Raikov and Kovaleva 1996). In

phological changes of aging. For example, the increasing contrast, macronuclei in other ciliates divide by amitosis for

staining darkness of macronuclei indicates DNA synthe- many generations until conjugation occurs. Macronuclear

sis (i.e. likely replication of certain genes; Bobyleva et al. amitosis is estimated to occur for 40–60 cell divisions in

1980; Kovaleva and Raikov 1978; Raikov 1994; Raikov et al. Tetrahymena thermophila (Class Oligomenophorea), and for

1963). DNA accumulation also generates macronuclei with more than 100 in Dileptus anser (Class Litostomatea; Lynn

a wide distribution of DNA content even within the same and Doerder 2012; Yudin and Uspenskaya 2007). The short

cell (Raikov 1994). For example, in Loxodes magnus, DNA life span of Div− macronuclei in Karyorelictea might func-

content of young macronuclei is ∼4C while aging macronu- tion as a mechanism to avoid the accumulation of deleterious

clei reach an average of 5–6C; in Kovalevaia sulcata, old mutations in Div− nuclei.

macronuclei vary in DNA content from 2C to 12C (Bobyleva

406 Y. Yan et al. / European Journal of Protistology 61 (2017) 399–408

Implications of unusual nuclear features in award 1R15GM113177 and NSF award 1541511to Dr. Laura

Karyorelictea A. Katz.

Since the 1950’s, karyorelictean nuclei have been investi-

gated primarily using cytochemical and microscopy methods, References

including Feulgen staining. Here we speculate on the impli-

cations of the patterns described above. We propose two

Al-Rasheid, K.A.S., 2001. New records of interstitial ciliates (Pro-

possible explanations for the maintenance of a minimum

tozoa Ciliophora) from the Saudi coasts of the Red Sea. Trop.

of two macronuclei within each cell. One possibility is Zool. 14, 133–156.

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We are grateful to Mr. Xyrus Maurer-Alcalá, Dr. Jean-

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David Grattepanche and the rest of the Katz lab for their

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C.T., Sebra, R.P., Landweber, L.F., 2014. The architecture of a

Thanks also to Dr. Weibo Song and Dr. John Clamp for their

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