European Journal of Phycology

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The host-range of Paraphysoderma sedebokerensis,

a chytrid that infects Haematococcus pluvialis

Jenia Gutman , Aliza Zarka & Sammy Boussiba

To cite this article: Jenia Gutman , Aliza Zarka & Sammy Boussiba (2009) The host-range of
Paraphysoderma�sedebokerensis, a chytrid that infects Haematococcus�pluvialis , European
Journal of Phycology, 44:4, 509-514, DOI: 10.1080/09670260903161024

To link to this article: https://doi.org/10.1080/09670260903161024

Published online: 11 Nov 2009.

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 Eur. J. Phycol., (2009), 44(4): 509–514

The host-range of Paraphysoderma sedebokerensis, a chytrid

that infects Haematococcus pluvialis

JENIA GUTMAN, ALIZA ZARKA AND SAMMY BOUSSIBA

The Landau Family Microalgal Biotechnology Laboratory, Jacob Blaustein Institutes for Desert Research, Ben-Gurion
University of the Negev, Sede Boker Campus 84990, Israel

(Received 19 January 2009; revised 27 May 2009; accepted 15 June 2009)

A parasitic chytrid that attacks the green alga Haematococcus pluvialis was recently isolated in our laboratory and identified as
a novel species from the phylum Blastocladiomycota, named herein Paraphysoderma sedebokerensis (nom. prov.). A method
for early and precise detection of chytrid infections was developed using the fluorescent dye, Nile red, which stained the
chytrids’ sporangia. Using this technique we determined the specificity of Paraphysoderma sedebokerensis for 13 algal species
belonging to the Chlorophyta. Algal species tested including: Chlamydomonas nivalis, Chlorella emersonii, Chlorella vulgaris,
Chlorococcum sp., Chlorogonium elongatum, Monoraphidium braunii, Muriella zofigiensis, Scenedesmus obliquus, Scenedesmus
vacuolatus (two strains), and Scotiellopsis oocystiformis were either resistant to infection, or only experienced slight levels of
infection during exponential growth. During exponential growth phase 100% Chlorella zofigiensis Donz cells were infected,
but none developed any infection during resting stage. Only in cultures of H. pluvialis did infections develop rapidly (3–4 days)
and intensively (100% cells infected) during both the logarithmic and stationary stages of growth. We suggest that the newly
isolated chytrid, Paraphysoderma sedebokerensis (nom. prov.), is highly specific for H. pluvialis, but has a limited capacity to
infect other green algae.

Key words: Blastocladiomycota, Chytrid, Haematococcus pluvialis, host-range, Nile red, Paraphysoderma sedebokerensis
(nom. prov.), pathogen, secondary carotenoids

Introduction alga Haematococcus pluvialis (Chlorophyta,

Microalgae are often plagued by harmful contami-
nants, including viruses, bacteria, protists, fungi,
and various grazers, both in their natural habitats
and in aquaculture. These parasites include zoos-
poric true fungi, or chytrids, which belong to the
fungal phylum Chytridiomycota (Canter-Lund &
Lund, 1995), or Blastocladiomycota (James et al.,
2006). Members of this class of fungi have been
found to cause extreme fluctuations in algal popu-
lations in natural fresh and marine water bodies
(Blinn & Button, 1973; Bruning et al., 1992).
Chytrid infections of algae have also been
described in sewage oxidation ponds (Abeliovich
& Dikbuck, 1977) and in outdoor algal mass-cul-
ture systems (Lukavsky, 1970). However, many
aspects of the ecology of chytrids still remain
poorly understood, including the nature of the
interactions between parasites and their hosts
(Gleason et al., 2008).
A novel parasite was recently isolated in our
laboratory (Hoffman et al., 2008). It was
found to contaminate cultures of the green
Correspondence to: Sammy Boussiba. E-mail: sammy@bgu.ac.il
ISSN 0967-0262 print/ISSN 1469-4433 online/09/040509–514 ß 2009 British Phycological Society
DOI: 10.1080/09670260903161024
Volvocales), the best natural source of the com-
mercially exploited ketocarotenoid, astaxanthin
(Olaizola, 2003). The life cycle of the parasite was
determined and its phylogeny characterized, based
on its 18S ribosomal DNA sequence data. These
indicated that it is a novel chytrid, closely related
to the vascular plant pathogen Physoderma
(Blastocladiomycota), therefore herein it has been
named Paraphysoderma sedebokerensis (nom.
prov.). In the initial stages of infection, only the
presence of an attached chytrid indicates contam-
ination. As the infection proceeds, the chytrid
sporangium grows bigger, as it degrades and con-
sumes the host cytoplasm and the algal remnants
turn into a brown, reduced, clumpy mass
(Hoffman et al., 2008).
Chytrid species exhibit considerable variation in
host ranges (Barr & Hickman, 1967a, b; Ibelings
et al., 2004). Canter & Jaworski (1982) observed
attraction of chytrid zoospores to a wide range of
hosts. However in contrast, narrow host specificity
based on species-specific interactions (Holfeld,
1998), or strain-specific interactions (De Bruin
et al., 2008) have also been demonstrated.

Published online 11 Nov 2009

J. Gutman et al.
In some cases extreme specificity for a particular
stage in the hosts life cycle has been reported
(Canter-Lund & Lund, 1995).
The chytrid that attacks H. pluvialis has been
shown to infect both the palmelloids (green cells)
and the aplanospores (red cyst). However, the fla-
gellates (young green cells released from ‘mother
cells’) were not susceptible to the chytrid infection
(Hoffman et al., 2008). In this study, the host-
range of the chytrid for different species of
green algae was tested using a newly developed
method based on staining employing Nile red flu-
orescent dye.
Materials and methods
Algal cultures
Most of the species were obtained from the Culture
Collection of Algae of the University of Göttingen
(SAG), Göttingen, Germany. Haematococcus pluvialis
Flotow 1844em.Wille K-0084 was obtained from the
Scandinavian Culture Collection of Algae and Protozoa
(SCCAP) at the University of Copenhagen, Denmark.
Chlorococcum sp. UTEX 819 and Chlamydomonas nivalis
UTEX 2824 were purchased from UTEX, the culture
collection of algae at the University of Texas, Austin.
Chlorella zofigiensis Donz CCAP 211/14 and Chlorella
emersonii CCAP 211/11N were obtained from the
Culture Collection of Algae and Protozoa (CCAP), at
the Scottish Association for Marine Science, UK.
Most of the species were grown in a modified BG11
medium (Boussiba & Vonshak, 1991). Chlorogonium
elongatum SAG 12-2b and Chlamydomonas nivalis
UTEX 2824 were grown in a basal medium
(Kobayashi et al., 1991).
Algal cultures at the logarithmic stage were obtained
as described in Boussiba & Vonshak (1991) to give a
H. pluvialis cell density of 2  105 cells ml1 after inocu-
lation. Other algae were inoculated at similar initial
chlorophyll content i.e. 5 mg chl ml1.
To obtain stressed cultures, algal cells harvested
from the logarithmic stage were re-suspended in
nitrogen depleted medium to give 1 mg ml1 chloro-
phyll, a concentration indicative for their optimum
exposure to illumination. Cultures (100 ml) in
250 ml Erlenmeyer flasks were then incubated on
rotary shaker for 10 days (temperature 25 C, illumi-
nation 80 mmol photon m2 s1, rotation speed
125 rpm).
Chytrid growth conditions
Chytrids were grown in chytrid growth medium (CGM)
(Hoffman et al., 2008), to which were added 1.25 g l1
yeast extract, 2.5 g l1 peptone and 5 g l1 glucose
(enriched CGM). This inoculum contained thriving
fungal cells (reaching OD of 3, within 3–5 days,
from initial OD of 0.025).
510
Chytrid inoculation of algal cultures
An axenic, 5-day-old chytrid culture at its logarithmic
stage (virulent toward Haematococcus), was used to
inoculate samples of either exponentially growing, or
stressed algal cultures. Cultures (20 ml) in 50 ml
Erlenmeyer flasks, or 5 ml/well in 6-well plates
(1  106 cells ml1) were inoculated with the chytrid cul-
ture to yield a final concentration of about 0.025
OD600 ml1. To ensure that all algal species were sub-
jected to identical experimental procedures, algal densi-
ties in the Erlenmeyer flasks and micro-well plates were
based on the equivalent dry weight of 106 H. pluvialis
cells ml1. The medium used for the experiments was
either mBG11, or mBG11 without nitrogen, to investigate
the susceptibility to infection of the logarithmic, or the
stressed stage, respectively. The inoculated cultures were
then incubated at 30 C, an optimal growth temperature
for the chytrid (Hoffman et al., 2008), on an orbital
shaker (125 rpm), under continuous dim white light
(15 mmol photon m2 s1). Under these experimental
conditions the pathogen is fully viable and the algae
can survive, but do not grow. The cultures were
observed daily over a period of seven days for the devel-
opment of contamination.
Nile red staining and epidemic determination
Pellets containing 2  105 cells were stained with 100–
200 mg Nile red l1 in 2% DMSO, mixed and washed
immediately with 1 ml DDW (10 s; 13,400 g). The pellet
was re-suspended in 20–40 ml DDW to obtain a dense
sample suitable for microscopic observation and mount-
ing. Samples were observed on a Axioskop1 (Zeiss)
microscope employing the light filter allowing maximum
exposure to the excitation wavelength (450–490 nm) and
a 520 nm cut-off filter.
Quantification of fungal infection was estimated by
determining the prevalence of infection, Pr (Bruning,
1991; Holfeld, 2000):
Pr ¼ Ni  100=ðNi þ Nu Þ
where Ni is the number of infected cells carrying spor-
angium and Nu is the number of uninfected cells. Each
test included 1500 algal cells screened. Experiments
were repeated three times and one representative exper-
iment is presented here.
Results
Nile-red staining of chytrid sporangia
To visualize chytrid sporangia mycologists use
Calcofluor white to stain chitin, the major constit-
uent in the chytrids’ cell walls (Kagami et al.,
2004). However, this dye also stains the cell wall
of H. pluvialis (Montsant et al., 2001), thus limiting
the ability to differentiate between chytrid
sporangia and the algal cells. We found that stain-
ing with the lipophilic dye Nile red (Greenspan
et al., 1985) facilitates the efficient detection of
Specificity of P. sedebokerensis infecting H. pluvialis
both, very young and mature, chytrid sporangia of
Paraphysoderma sedebokerensis (nom. prov.)
(Figs 1–4). Staining was evident as yellow/gold flu-
orescence (emission at 4520 nm), under blue exci-
tation (450–490 nm). This new, simple staining
method does not require fixation and there is no
need for prolonged incubation with the dye. In
fact, the threshold of detection of the colourless
sporangia, at up to 1% of infected cells without
staining, is remarkably reduced upon Nile red
staining, (Figs 5, 6). Furthermore the sensitivity
of the technique allows detection of young sporan-
gia even in dense algal cultures.
Chytrid specificity towards different microalgae
Thirteen algal species, including representatives
of the major/most common Chlorophyte genera,
were tested for their susceptibility to infection by
Paraphysoderma sedebokerensis (nom. prov.).
The susceptibility test was conducted at 30 C,
a temperature optimal for the growth of the
Figs 1–6. Infected Haematococcus pluvialis culture stained
with Nile red. Images are either in phase contrast (1, 3 and
5), or under fluorescent light (2, 4 and 6). Figs 1, 2: Nile-red
staining enables very young sporangium to be detected.
Figs 3, 4: lipid globules inside the chytrid’s sporangium.
Figs 5, 6: Staining of infected culture allows for the algal
cells carrying fungal sporangia to be counted. Arrows indi-
cate the location of a chytrid sporangium, which is unde-
tectable under phase contrast in dense culture.
511
chytrid, to facilitate epidemic development
(Hoffman et al., 2008). During the 7 days of exper-
iment the algae did not grow, but remained viable,
unless they were infected by the chytrid. The kinet-
ics of infection development in the algal species
that were infected at their logarithmic stage is
depicted in Fig. 7. Epidemics developed rapidly
and intensively only in H. pluvialis and C. zofigien-
sis Donz, i.e. after 3–4 days almost all the cells were
infected and their contents depleted. Three addi-
tional species – S. vacuolatus (SAG 211-8b); M.
zofigiensis and S. oocystiformis – served as hosts
for the pathogen during their logarithmic growth
stages, although the infection developed at a slower
rate and did not affect more than 20% of the algal
cells, even after 7 days of exposure. The remaining
eight species, including other species of Chlorella
and Scenedesmus, were not infected at all (data not
shown). After 7 days incubation, cultures of
H. pluvialis and C. zofigiensis Donz collapsed
and turned brown in colour. Since a 7-day incuba-
tion period was sufficiently to result in complete
‘collapse’ of H. pluvialis cultures the prevalence
of infection in the other algal species after 7 days
was used as measure for characterization of strain
susceptibility (Table 1).
The specific attachment of zoospores onto a par-
ticular host, or group of algal species, indicates
that specific signals are involved in the process
(Kagami et al., 2007). To test whether these signals
were preserved in algae after they were exposed to
stress conditions, we performed an additional sus-
ceptibility test on the different species after they
100
Prevalence of infection (%)
80
60
40
20
0
0 1 2 3 4 5 6 7
Days from inoculation
Fig. 7. Development of ‘epidemics’ in different algal species
at their logarithmic stage, during the period of 7 days. Each
point on the graph represents the percentage of algal cells
observed carrying Paraphysoderma sedebokerensis (nom.
prov.) sporangium. Haematococcus pluvialis Flotow
1844em.Wille SCCAP K-0084 (.), Chlorella zofigiensis
Donz CCAP 211/14 (#), Scenedesmus vacuolatus
SAG 211-8b(*), Muriella zofigiensis SAG 4.80 (m),
Scotiellopsis oocystiformis SAG 277-1(¨).
J. Gutman et al. 512
Table 1. A survey of the susceptibility of algal species to defined as susceptible, a zoospore must be able
infection by P. sedebokerensis in vegetative and nitrogen-
not only to attach to, but also to penetrate, the
starved stages.
algal cell wall. Furthermore, the zoospore must
Development of epidemic
utilize the cellular constituents of the alga to
grow and proliferate. This definition coincides
Vegetative with the earlier suggestion of Holfeld (1998) that
Algal strain stage N-starved host specificity should be expressed during encyst-
Haematococcus pluvialis Flotow 100 100 ment of the zoospore rather than at the earlier
1844em.Wille SCCAP K-0084 stages of possible recognition.
Chlorella zofigiensis Donz CCAP 211/14 100 0 The Nile-red staining protocol, as described in
Chlorella emersonii CCAP 211/11N 1 0 the present work, allows efficient and sensitive
Chlorella vulgaris SAG 211-8L 0.4 0
Scenedesmus vacuolatus SAG 211-8b 17 0 detection of the chytrid sporangia of H. pluvialis –
Scenedesmus vacuolatus SAG 211-15 0.4 0 Paraphysoderma sedebokerensis (nom. prov.). It
Scenedesmus obliquus SAG 276-3a 0.4 0 might be also a useful tool for the detection of chy-
Muriella zofigiensis SAG 4.80 8 0 trids in algal culture, or in field, since these patho-
Chlorococcum sp. UTEX 819 0.8 0
Chlamydomonas nivalis UTEX 2824 0.1 0 gens are often undetected by non-specialists, or
Chlorogonium elongatum SAG 12-2b 0 0 confused with other organisms (Kagami et al.,
Monoraphidium braunii SAG 202-7b 0 0 2007). This is of particular interest in biotechnolo-
Scotiellopsis oocystiformis SAG 277-1 3 0 gical applications, since in some cases early detec-
tion of the contaminant is crucial to the survival of
Extent of fungal abundance in the culture was measured daily up to
7 days. The occurrence of adhesion and the estimated extent of the the culture, especially when the contaminant
epidemic – prevalence of infection, Pr – after 7 days are develops epidemics rapidly. Although Nile red has
represented. previously been used to visualize chytrid zoospores
(Kagami et al., 2004), the protocol described in this
were exposed to stress conditions (N starvation). study, for staining chytrid sporangia on algae, has a
To maintain the different algae in the same physi- particular advantage for phycologists that most fre-
ological state, this susceptibility test was performed quently detect a contaminant on algae and need to
in the absence of nitrogen. However, nitrogen by identify it. Under the blue excitation, as reported in
itself has no effect on chytrid virulence, at least this work the neutral lipids, rather than polar lipid
when tested in cultures of H. pluvialis (Hoffman are stained (Greenspan, 1985), emitting a charac-
et al., 2008). Excluding C. vulgaris, all species teristic yellow–gold fluorescence clearly distin-
exposed to N depletion accumulated secondary guishable from the red chlorophyll fluorescence of
carotenoids, including Chlorogonium elongatum, the algae (see Figs 1–6). Moreover, the staining
S. obliquus and M. braunii, all three of which does not require sample preparation or incubation
have not previously been reported as secondary time with the dye.
carotenoid producers (data not shown). However, Of all the algae tested H. pluvialis provided the
of the N-starved species, only H. pluvialis was sus- best conditions for chytrid adhesion and prolifera-
ceptible to chytrid infection (Table 1). The kinetics tion, as indicated by sporangia development.
of the infection and epidemic development in cul- Haematococcus pluvialis is highly susceptible to
tures of this alga after nitrogen starvation were the chytrid both at the logarithmic and cyst
comparable to the effects observed in the green (nitrogen-starved) stages of the alga. In contrary,
vegetative culture (3–4 days). In all the other spe- other susceptible species – C. zofigiensis Donz,
cies examined, even 7 days after inoculation with S. vacuolatus SAG 211-8b, M. zofigiensis, and
the chytrid, they were not infected by the chytrid S. oocystiformis – were attacked by the chytrid
and no change in their cell density was observed. only during logarithmic growth. Furthermore, the
These results indicate that H. pluvialis is the ‘best’ latter three species did not develop epidemics.
host for the chytrid as infection and development On the basis of this observation we suggest that
of the epidemic occurred fastest in the logarithmic P. sedebokerensis (nom. prov.) is highly specific
and the N-starved H. pluvialis cultures (Table 1). for H. pluvialis.
The speed with which the epidemic developed in The narrow host range of the chytrid is high-
the cultures – within 3 days of inoculation – indi- lighted when comparing the susceptibilities of
cated that H. pluvialis was the most susceptible different species of the same algal genus. For exam-
algal species tested. ple, of the different Chlorella species, C. zofigiensis
Donz was ‘recognised’ by the chytrid during
the alga’s logarithmic stage, while the other two
Discussion
species, C. emersonii and C. vulgaris, were not
This study defines host specificity on the basis of part of the chytrid host range. Among the species
epidemic development and for an alga to be of Scenedesmus studied, only S. vacuolatus
Specificity of P. sedebokerensis infecting H. pluvialis
SAG 211-8b exhibited ongoing recognition, as
the other two, S. vacuolatus SAG 211-15 and S.
obliquus SAG 276-3a, were apparently not suscep-
tible to infection by the pathogen. These observa-
tions are in agreement with those of Gromov
(1999), who found that the sensitivity of
Chlorococcalean algae to a given strain of chytrid
was strain-specific, but not genus or even species
specific. However, P. sedebokerensis (nom. prov.)
appears to be genus-specific to the Haematococcus
genus. Cultures of various strains were readily
infected as described in a previous study: H. lacus-
tris NIES-144, H. lacustris UTEX 16, H. pluvialis
SAG 192.8, and H. lacustris CCAP 34/19 (Hoffman
et al., 2008) and an additional 15 strains were sus-
ceptible to the pathogen in their palmelloid stage.
The chytrid parasite studied here is specific to
the Haematococcus genus, both at the palmelloid
and the cyst stages and rarely recognizes, or infects,
other algal species. One possible explanation for
the chytrid susceptibility of H. pluvialis, also
during its cyst stage, is that the changes in the
cell wall that H. pluvialis undergoes upon encyst-
ment are quantitative and not qualitative, such
that the cell wall components exploited by the
parasite – for recognition and adsorption – are
not changed by the entrance of H. pluvialis into
its cyst stage (Montsant et al., 2001).
As suggested by Coder & Goff (1986), narrow
host specificity may be related to cell surface prop-
erties. One of these properties was suggested to be
the sporopollenin fraction found in the cell wall,
but not as a single mean. The restriction Coder and
Goff put on this component as a single means of
protection correlates with our results. Likewise,
species that do not contain algaenan (sporopol-
lenin from an algal source), such as
Chlorococcum sp. UTEX 819 (Gelin et al., 1997)
and C. vulgaris SAG 211-8L (Burczyk et al., 1999),
are apparently resistant to chytrid attack, unlike
species that are reported to contain that biomacro-
molecule, i.e., H. pluvialis (Montsant et al., 2001)
and S. vacuolatus SAG 211-8b (Derenne et al.,
1992). The fact that H. pluvialis cell wall contains
algaenan and still is penetrated/digested by the
chytrid fungus is remarkable. Work is underway
to clarify this unusual host range specificity of
P. sedebokerensis (nom. prov.) to its prey the
green alga H. pluvialis.
References
ABELIOVICH, A. & DIKBUCK, S. (1977). Factors affecting infection of
Scenedesmus obliquus by a Chytridium sp in sewage oxidation
ponds. Appl. Environ. Microbiol., 34: 832–836.
BARR, D.J.S. & HICKMAN, C.J. (1967a). Chytrids and algae I. Host-
substrate range and morphological variation of species of
Rhizophydium. Can. J. Bot., 45: 423–430.
513
BARR, D.J.S. & HICKMAN, C.J. (1967b). Chytrids and algae II.
Factors influencing parasitism of Rhizophydium sphaerocarpum
on Spirogyra. Can. J. Bot., 45: 431–440.
BLINN, D.W. & BUTTON, K.S. (1973). Effect of temperature on
parasitism of Pandorina-sp by Dangeardia mammillata Schröd
in an Arizona mountain lake. J. Phycol., 9: 323–326.
BOUSSIBA, S. & VONSHAK, A. (1991). Astaxanthin accumulation in
the green-alga Haematococcus pluvialis. Plant Cell Physiol., 32:
1077–1082.
BRUNING, K. (1991). Effects of phosphorus limitation on the epi-
demiology of a chytrid phytoplankton parasite. Freshwater Biol.,
25: 409–17.
BRUNING, K., LINGEMAN, R. & RINGELBERG, J. (1992). Estimating
the impact of fungal parasites on phytoplankton populations.
Limnol. Oceanogr., 37: 252–260.
BURCZYK, J., SMIETANA, B., TERMINSKA-PABIS, K., ZYCH, M. &
KOWALOWSKI, P. (1999). Comparison of nitrogen content
amino acid composition and glucosamine content of cell
walls of various chlorococcalean algae. Phytochemistry, 51:
491–497.
CANTER, H.M. & JAWORSKI, G.H.M. (1982). Some observations on
the alga Fragilaria crotonensis Kitton and its parasitism by two
chytridiaceous fungi. Am. Bot., 49: 429–446.
CANTER-LUND, H. & LUND, J.W.G. (1995). Freshwater Algae: Their
Microscopic World Explored. Bristol, UK: Biopress.
CODER, D.M. & GOFF, L.J. (1986). The host range of the chlorel-
lavorous bacterium (Vampirovibrio chlorellavorus). J. Phycol., 22:
543–546.
DE BRUIN, A.R., IBELINGS, B.W., KAGAMI, M., MOOIJ, W.M. &
VAN DONK, E. (2008). Adaptation of the fungal parasite
Zygorhizidium planktonicum during 200 generations of
growth on homogeneous and heterogeneous populations of
its host, the diatom Asterionella formosa. Euk. Microbiol., 55:
69–74.
DERENNE, S., LARGEAU, C., BERKALOFF, C., ROUSSEAU, B.,
WILHELM, C. & HATCHER, P.G. (1992). Nonhydrolyzable macro-
molecular constituents from outer walls of Chlorella fusca and
Nanochlorum-eucaryotum. Phytochem., 31: 1923–1929.
GELIN, F., VOLKMAN, J.K., DE LEEUW, J.W. & DAMSTE, J.S.S.
(1997). Mid-chain hydroxy long-chain fatty acids in microalgae
from the genus Nannochloropsis. Phytochem., 45: 641–646.
GLEASON, F., KAGAMI, M., LEFEVRE, E. & SIME-NGANDOS, T.
(2008). The ecology of chytrids in aquatic ecosystems: Roles in
food web dynamics. Fungal Biol. Rev., 22: 17–25.
GREENSPAN, P., MAYER, E.P. & FOWLER, S.D. (1985). Nile red: a
selective stain for intracellular lipid droplets. J. Cell Biol., 100:
965–73.
GROMOV, B., PLJUSCH, A. & MAMKAYEVA, K. (1999). Cultures of
Rhizophydium spp. (Chytridiales) parasites of chlorococalean
algae. Algol. Stud., 95: 115–123.
HOFFMAN, Y., AFLALO, C., ZARKA, A., GUTMAN, J., JAMES, T.Y. &
BOUSSIBA, S. (2008). Isolation and characterization of a
novel pathogenic chytrid species from the phylum
Blastocladiomycota, parasitic on the green alga Haematococcus.
Mycol. Res., 112: 70–81.
HOLFELD, H. (1998). Fungal infections of the phytoplankton:
Seasonality, minimal host density, and specificity in a meso-
trophic lake. New Phytol., 138: 507–517.
HOLFELD, H. (2000). Infection of the single-celled diatom
Stephanodiscus alpinus by the chytrid Zygorhizidium: parasite
distribution within host population, changes in host cell size,
and host-parasite size relationship. Limnol. Oceangr., 45:
1440–1444.
IBELINGS, B.W., DE BRUIN, A., KAGAMI, M., RIJKEBOER, M.,
BREHM, M. & VAN DONK, E. (2004). Host parasite interactions
between freshwater phytoplankton and chytrid fungi
(Chytridiomycota). J. Phycol., 40: 437–453.
JAMES, T.Y., LETCHER, P.M., LONGCORE, J.E., MOZLEY-
STANDRIDGE, S.E., PORTER, D., POWELL, M.J., GRIFFITH, G.W.
& VILGALYS, R. (2006). A molecular phylogeny of the flagellated
fungi (Chytridiomycota) and description of a new phylum
(Blastocladiomycota). Mycologia, 98: 860–871.
J. Gutman et al.
KAGAMI, M., DE BRUIN, A., IBELINGS, B.W. & VAN DONK, E.
(2007). Parasitic chytrids: their effects on phytoplancton commu-
nities and food-web dynamics. Hydrobiologia., 578: 113–129.
KAGAMI, M., VAN DONK, E., DE BRUIN, A., RIJKEBOER, M. &
IBELINGS, B.W. (2004). Daphnia can protect diatoms from
fungal parasitism. Limnol. Oceanogr., 49: 680–685.
KOBAYASHI, M., KAKIZONO, T. & NAGAI, S. (1991). Astaxanthin
production by a green-alga, Haematococcus pluvialis accompa-
nied with morphological-changes in acetate media. J. Ferment.
Bioeng., 71: 335–339.
514
LUKAVSKY, J. (1970). Phlyctidium scenedesmi, a chytrid destroying
an outdoor mass culture of Scenedesmus obliqus. Nova Hedwig.,
19: 775–777.
MONTSANT, A., ZARKA, A. & BOUSSIBA, S. (2001). Presence of a
nonhydrolyzable biopolymer in the cell wall of vegetative cells
and astaxanthin-rich cysts of Haematococcus pluvialis (chloro-
phyceae). Mar. Biotechnol., 3: 515–521.
OLAIZOLA, M. (2003). Commercial development of microalgal bio-
technology: From the test tube to the marketplace. Biomol. Eng.,
20: 459–466.