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oral apparatus is dedifferentiated as the proter becomes the dispersal \u201c swarmer \u201d stage, which has a very reduced spiral of membranelles and does not form food vacuoles. Differentiation of the proter oral apparatus occurs upon settling and shows a similar pattern to the development of the opisthe oral apparatus prior to cell division (Mulisch & Hausmann, 1988). Heterotrichs, like the karyorelicteans, have remarkable regenerative abilities that have long been exploited to probe how the development of cell pattern is regulated at the morphological (e.g., De Terra, 1985; Fahrni, 1985; Tartar, 1961; Uhlig, 1960) and biochemical levels (Bohatier, 1981, 1995), a subject area that is well beyond the scope of our treatment here, but see Frankel (1989) for a comprehensive review. 6.6 Nuclei, Sexuality and Life Cycle The macronuclei of heterotrichs range from compact ellipsoid to ribbon-like and finally to moniliform or beaded. Correlated with their large cell size, heterotrich macronuclei are large and highly amplified. Nucleoli are often promi- nent. In Stentor , the nuclear envelopes of both macronucleus and micronuclei are surrounded by an additional membrane system that integrates them structurally to the endoplasmic reticulum (Mulisch, 1988). Since Mulisch (1988) used special fixation techniques to obtain this result, it may be a more general property of heterotrichs than is presently realized. Heterotrichs typi- cally have many micronuclei distributed along the length of filiform macronuclei or associated singly or in groups with each bead of moniliform macronuclei. During cell division, filiform and moniliform macronuclei become compact and nucleoli dedif- ferentiate. Macronuclear division is accomplished primarily by extramacronuclear microtubules (Diener et al., 1983; Jenkins, 1973). These extrama- cronuclear microtubules may be intimately associ- ated with the cortex since De Terra (1983) has demonstrated cortical control over the direction of macronuclear elongation in Stentor . Since the majority of ciliates use intramacronuclear microtu- bules in macronuclear karyokinesis , Orias (1991a) has argued that extramacronuclear microtubules represent an independent evolution of the capacity to divide the macronucleus (see Chapter 4 ). Relatively little research has been done on the molecular biology of heterotrich nuclei. The macro- nuclear DNA molecules or \u201cchromosomes\u201d are typically long, some being up to 20 µm (Hufschmid, 1983; Pelvat & De Haller, 1976). Thus, very little frag- mentation of micronuclear chromosomes appears to occur, in contrast to other classes (Riley & Katz, 2001; Steinbrück, 1990). The heterotrich Blepharisma at least shows a deviation from the use of the universal stop codons \u2013 UAA, UGA, and UAG. Of the three, it uses at least UAA, like the spirotrich Euplotes (Liang & Heckmann, 1993) (see Chapter 7 ). As noted above (see Life History and Ecology ), conjugation may be initially stimulated by starvation conditions or changes in temperature. These condi- tions stimulate transcription of a gamone gene (Sugiura, Kawahara, Iio, & Harumoto, 2005), which is followed by excretion of diffusible mating type substances, called gamones (Miyake, 1996) or mating pheromones (Luporini & Miceli, 1986). All species of Blepharisma excrete two gamones: blepharismone , a tryptophan derivative resembling serotonin (Kubota, Tokoroyama,Tsukuda, Koyama, & Miyake, 1973; Miyake & Bleyman, 1976) com- mon to all species; and blepharmone , a 20-kDa glycoprotein that is species specific (Braun & Miyake, 1975). Typically, the diffusion of these substances is sufficient to stimulate conjugation in receptive individuals of fresh-water species of Blepharisma but is apparently not sufficient in marine species (Ricci & Esposito, 1981). There is considerable debate about the precise mechanisms that stimulate conjugation in hetero- trichs at the cellular and molecular levels. Miyake (1996) favors his gamone-receptor hypothesis while Luporini and Miceli (1986) reinterpret the results from Blepharisma in the context of their self-recogni- tion hypothesis . Whichever interpretation is true, the heterotrichs have not evolved a stable mating type system, as stable lines are quite rare (Demar-Gervais, 1971; Miyake & Harumoto, 1990). Luporini and Miceli (1986) argued that Blepharisma (and therefore heterotrichs in general?) has not yet evolved a mating type system and that it is only the formation of blep- harmone-blepharismone complexes that stimulate conjugation . Once conjugation is stimulated and micronuclear meiosis has occurred, a variable number of micro- nuclei enter the third or pregametic division: one in Spirostomum and Blepharisma americanum , but two or three in Fabrea and Blepharisma japonicum (Raikov, 1972). Ultimately, the products of only one of these micronuclei form the stationary and migratory gametic nuclei, which fuse to form the synkaryon . Thus, the exconjugants are genetically identical to each other although different from their parents. The synkaryon typically divides three times to produce eight products, several of which develop as macronuclear anlagen that may fuse to form the macronucleus, following separation of the conjugants (Raikov, 1972). 6.7 Other Features Given the widespread distribution of ciliates, and heterotrichs in particular, and their large cell size and ease of cultivation, several laboratories have developed low-cost bioassays or microbiotests using Spirostomum species. Spirostomum turns out to be quite a sensitive indicator species, especially to some heavy metal contaminants (Madoni, 2000; Nalecz-Jawecki, 2004; Nalecz-Jawecki & Sawicki, 2002; Twagilimana, Bohatier, Grolière, Bonnemoy, & Sargos, 1998). 6.7 Other Features 139