Abstract

A cluster of antigenic, genomic, karyotypic, isoenzymatic and morphological differences have been reported among Pneumocystis populations. Multilocus enzyme electrophoresis revealed strong linkage disequilibrium suggesting that Pneumocystis genotypes from different hosts have been genetically isolated from each other for a very long time. At least in some cases, genetic diversity is associated with phenotypic differences as revealed by in vitro, ultrastructural and cross infection studies. Thus, biodiversity in Pneumocystis has obvious epidemiological implications. Cross infection experiments revealed that Pneumocystis host species-related genetic differences are associated with close host species specificity, which suggests that transmission cannot take place between hosts of different species and that immunocompromised patients contract the infection primarily from infected humans. But these affirmations do not preclude other reservoirs for human pneumocystosis and research has to be extended to natural populations of synanthropic or wild mammals. Transmission of human pneumocystosis was also approached by typing human Pneumocystis isolates from patients or carriers, which should allow the follow up of parasite strains in human populations. As the strains of Pneumocystis found in different host species were considered for a long time to be morphologically indistinguishable, only one species of Pneumocystis was accepted for almost one century. At present, the scientific community is progressively accepting that the terminology ‘P. carinii’ is hiding a heterogeneous group of microorganisms. As available data made it impossible to establish if genetic divergence derives from clonal reproduction or speciation, no new species names have been attributed to Pneumocystis populations, but a trinomial nomenclature, including the Latin name of the host, was adopted in 1994. It has to be outlined finally that works on biodiversity of Pneumocystis populations are basically important as they have revealed a new group of eukaryotic, pathogenic, heterogeneous microorganisms with fungal affinities, difficult to cultivate until now and widely spread in ecosystems. These researches are opening a virgin field for microbiology research.

Introduction

Pneumocystis biodiversity was already reported in the past by Frenkel [1]. This author observed host specificity and antigen differences between rat and human Pneumocystis, and thus considered human Pneumocystis a different species from that of rat Pneumocystis. He proposed the term ‘P. jiroveci’ for human Pneumocystis, in honor to Dr. Otto Jirovec who recognized Pneumocystis as the agent of interstitial plasma cell pneumonia. Nevertheless, his proposition was usually disregarded by the scientific community. Recently, a cluster of antigenic [2–6], genomic [7–11], karyotypic [10, 12], isoenzymatic [13–15] and morphological differences [13] have been reported among Pneumocystis populations.

The majority of differences among parasite populations were reported on nuclear and mitochondrial DNA, and the most frequently employed marker was a sequence of the mitochondrial large subunit ribosomal RNA gene (mt LSU rDNA). Moreover, few studies were done on sequences encoding proteins interacting with sulfa drugs, for example dihydropteroate synthase (DHPS). The nucleotide sequence of this sulfa target enzyme differed substantially in human-, rat- and mouse-derived Pneumocystis[16]. Moreover, six nucleotide changes were found in six human isolates.

On the whole, analyzed Pneumocystis nucleic acids revealed differences in terms of sequence, but also in terms of length and presence of introns. Thus, a pattern has emerged whereby there appeared to be three levels of divergence termed: (i) class I, seen between isolates of Pneumocystis from the same host species; (ii) class II, observed between laboratory rodent isolates of Pneumocystis (two forms found in the rat, and one found in the mouse); (iii) class III, observed between isolates from different host species ([17]; see Chapter I of this issue). Recently, preliminary data obtained from additional isolates of Pneumocystis harvested from pigs and ferrets, suggested that the situation may be more complex (see chapter I).

The genetic diversity of Pneumocystis populations has also been investigated using another tool, multilocus enzyme electrophoresis (MLEE) [13–15], allowing genetic population studies. Analyzing 70 parasite isolates from rats, mice and rabbits, with five enzyme systems, only five different multilocus associations (zymodemes) were recorded. These results corroborate molecular studies. Furthermore, isoenzyme analysis strongly suggests that different hosts harbor drastically distinct categories of Pneumocystis genotypes. Moreover, this considerable level of linkage disequilibrium suggests that Pneumocystis genotypes from different hosts have been genetically isolated from each other for a very long time, i.e. that they have undergone prolonged genetic and functional adaptation to each mammalian species.

Some differences in the in vitro behavior were reported among Pneumocystis isolates from rat and mouse [18]. Thus, rat-derived Pneumocystis organisms seem to have a higher capacity for attaching in vitro to target cells than mouse-derived parasites ([18] and unpublished observations). Moreover, the in vitro attachment of rat Pneumocystis seemed more sensitive to pentamidine or cytochalasin B than that of mouse-derived organisms [18]. Furthermore, a few but neat ultrastructural differences between rabbit and mouse Pneumocystis were found [13, 19]. Filopodia are much more thin, numerous and dendritic in parasites from mouse than in those from rabbit (Fig. 1).

Figure 1

Ultrastructural differences between rabbit-derived and mouse-derived Pneumocystis organisms. On the left, the two figures (top and bottom) show rabbit-derived Pneumocystis trophozoites with typical thick filopodia (arrowheads), attached to the lung epithelium (arrow). On the right, the two figures show mouse-derived Pneumocystis: several attached trophozoites (top) display numerous, thin, dendritic filopodia (arrowheads); an attached mature cyst (bottom) is separated from the epithelium by a space filled by abundant, thin filopodia and by a tangentially sectioned flattened Pneumocystis cyst. Magnification: left (top): ×9363; left (bottom): ×11 369; right (top): ×4125; right (bottom): ×7180.

Figure 1

Ultrastructural differences between rabbit-derived and mouse-derived Pneumocystis organisms. On the left, the two figures (top and bottom) show rabbit-derived Pneumocystis trophozoites with typical thick filopodia (arrowheads), attached to the lung epithelium (arrow). On the right, the two figures show mouse-derived Pneumocystis: several attached trophozoites (top) display numerous, thin, dendritic filopodia (arrowheads); an attached mature cyst (bottom) is separated from the epithelium by a space filled by abundant, thin filopodia and by a tangentially sectioned flattened Pneumocystis cyst. Magnification: left (top): ×9363; left (bottom): ×11 369; right (top): ×4125; right (bottom): ×7180.

At least on the basis of genomic, antigenic, isoenzymatic and ultrastructural data, the Pneumocystis biodiversity is correlated with the host species. These data came essentially from studies performed on Pneumocystis isolates from humans and laboratory animals (rats, mice, ferrets and rabbits). Only some studies were conducted on domestic animals like foals or pigs [3, 20] or on wild mammals (rodents, insectivorous, lagomorphous) [21–25].

The epidemiological impact of the Pneumocystis genetic diversity

The genetic biodiversity of Pneumocystis has obvious epidemiological implications in terms of host species specificity, P. carinii pneumonia (PCP) transmission and, potentially, in terms of virulence or drug resistance.

First, the host species-related parasite genetic diversity raises the notion of close host species specificity (stenoxenism), i.e. PCP transmission only between hosts of a same species. In order to investigate Pneumocystis host specificity, cross infection experiments were performed in our lab by using SCID mice or nude rats as experimental hosts. Inocula were parasite isolates from rat, mouse or rabbit. Only mouse-derived Pneumocystis developed in SCID mice and only rat-derived Pneumocystis developed in nude rats [26, 27]. The total absence of parasites in hosts inoculated with organisms from another host species was verified using PCR and hybridization. These molecular methods were also used in order to identify accurately the parasite strains with DNA probes specific for Pneumocystis from the three mammals. Likewise, ferret- or monkey-derived Pneumocystis were not infectious to the SCID mouse [28, 29]. These experiments revealed that host species-related genetic differences among Pneumocystis strains are associated with close host species specificity.

Thus, host species-related differences of Pneumocystis strains are associated with a marked host species specificity. This fact could have obvious implications on pneumocystosis transmission. Indeed, it suggests that (i) animal-to-animal transmission cannot take place between hosts of different species, (ii) human PCP is not a zoonotic disease, and (iii) immunocompromised patients contract PCP primarily from infected humans [30]. Of course, these affirmations do not preclude other reservoirs for human pneumocystosis, as mentioned by Wakefield [31]. Actually, man could not be the sole reservoir for the human PCP agent. In other pathogenic fungal groups, a variable host species specificity exists among closely related species. This is particularly clear in Dermatophytes, where four levels of host specificity may be distinguished, even within one genus [32]. Thus, the fact that rabbit-, rat-, mouse- or rabbit-derived Pneumocystis have been found to be stenoxenic does not means that P. carinii hominis (=P. carinii sp.f. hominis) is unable to grow in the lung of other mammals. In other words, human Pneumocystis strains could be oligoxenic or even euryxenic [33].

Moreover, research on Pneumocystis diversity has to be extended to natural populations of synanthropic or wild mammals, in order to avoid difficulties derived from conventional breeding. Actually, as most data supporting the hypothesis of the host species specificity of Pneumocystis resulted from laboratory animal studies, it could be speculated that these hosts, even from different strains, could come from a limited number of original standard colonies infected by a small number of Pneumocystis strains. This possibility has to be considered all the more since we know that the infection is easily transmitted by the airborne route [34] or in rabbits, from mothers to fetuses through the placenta [35]. Thus, identifying potential reservoirs for the agent of the human PCP, searching for Pneumocystis organisms in domestic, synanthropic or wild mammals as well as characterizing genomically the parasites found in these hosts remain essential objectives.

The transmission of human pneumocystosis can also be approached by typing human Pneumocystis isolates from patients [17, 36] or carriers [37]. Actually, typing should allow the follow up of parasite strains in human populations. Tsolaki and his collaborators [36] have shown that recurrent PCP can be caused by the same parasite strain when recurrence occurred within 8 months. Moreover, they reported that a predominant parasite strain may be responsible for recurrent episodes of PCP.

No data are available on drug resistance in Pneumocystis. However, several nucleotide changes have been found in highly conserved regions of the human Pneumocystis DHPS gene and they are similar to those responsible for sulfa resistance in other organisms [16]. The authors proposed two hypotheses to explain these changes: (i) they result from positive selection, which is consistent with selection pressure deriving from frequent exposure of parasites to sulfa drugs; (ii) Pneumocystis that infect humans would be a mixed population of parasites with different DHPS nucleotide sequences. From a practical point of view, it has to be underlined that we are able to identify genetically human Pneumocystis isolates but we do not yet have reliable in vitro systems allowing us to isolate parasite samples from patients, in order to measure their drug susceptibility.

Taxonomic impact

As parasite strains found in different host species were morphologically indistinguishable for a long time, P. carinii was considered for almost one century a widely spread, euryxenic, unique species. At present, this concept is changing and the scientific community is progressively accepting that the terminology ‘P. carinii’ is hiding a heterogeneous group of microorganisms with marked fungal affinities [35, 38].

The marked host species-related Pneumocystis diversity, supported now by genotypic and phenotypic evidence, leads to the question if Pneumocystis strains from different mammals have to be raised to the species rank [38], according to the biological concept, “species are groups of interbreeding natural populations that are reproductively isolated from other such groups„[39]. Therefore, populations of a true species must be unable to mate productively with populations of any other species. In fact, the attribution of the species rank to different populations of closely related organisms depends on the absence of genetic exchange among them.

Population genetics has provided useful tools for monitoring genetic flows among populations. In the case of microorganisms, population genetics methods rely mainly on the analysis of linkage disequilibrium (=nonrandom association of genotypes occurring at different loci), which requires the survey of several loci, and cannot therefore be completed by PCR amplification of isolated genes. Indeed, if several primers are used on a same isolate, and if this isolate is composed of several genotypes (mixed stock), it is not sure whether different primers will amplify the same genotype. Moreover, RAPD methods [14] are not useable in the case of Pneumocystis in so far as no continuous cultures are available for these organisms; actually, contaminating host DNA is constantly present in parasite samples. For these reasons, MLEE is at present the only method available for the joint analysis of several loci in Pneumocystis.

Using MLEE, Mazars et al. [14] have clearly shown that Pneumocystis isolates from three laboratory mammals (rabbit, rat and mouse) represent distinct genotypes and that these genotypes are genetically isolated from each other. All linkage disequilibrium tests have shown a high deviation from panmictic expectation, i.e. a strong nonrandom association between genotypes at different loci. These observations, which are consistent with phylogenetic analyses developed on MLEE or molecular data performed by many teams [40], strongly suggest that no genetic exchange occurs between Pneumocystis genotypes from different host species. Moreover, as Pneumocystis organisms isolated from a given mammal are apparently unable to grow in any other host species, there may be few opportunities for mating between parasites found in different mammal species [38].

Nevertheless, the scientific community has considered that additional evidence of no genetic exchange between the parasite populations from different host species is needed before considering them different Pneumocystis species. Therefore, giving them the status of distinct species remains a matter of convenience at present [14]. In this way, the Pneumocystis International Workshop of Cleveland [17] considered that in so far as available data made it impossible to distinguish genetic divergence derived from clonal reproduction from that expressing speciation, no new species names should be attributed to the Pneumocystis organisms from different host species. However, considering the marked host species-related biodiversity of Pneumocystis populations, a trinomial nomenclature, which includes the Latin name of the host species (e.g. P. carinii sp.f. hominis, P. carinii sp.f. rattus, P. carinii sp.f. oryctolagi, where sp.f.=special form or forma specialis) was adopted [17, 38].

Finally, works revealing a marked biodiversity in Pneumocystis organisms are basically important because they have opened up a virgin field for research: they have revealed a new group of eukaryotic, pathogenic, heterogeneous microorganisms with fungal affinities, difficult to cultivate up to date, widely spread in ecosystems [33, 41–43].

References

[1]
Frenkel
J.K.
(
1976
)
Pneumocystis jiroveci n.sp. from man: morphology, physiology and immunology in relation to pathology
.
Natl. Cancer Inst. Monogr.
 
43
,
13
30
.
[2]
Bauer
N.L.
Paulsrud
J.R.
Bartlett
M.S.
Smith
J.W.
Wilde
C.E.
(
1993
)
Pneumocystis carinii organisms obtained from rats, ferrets and mice are antigenically different
.
Infect. Immun.
 
61
,
1315
1319
.
[3]
Christensen
C.B.V.
Settnes
O.P.
Bille-Hansen
V.
Jorsal
S.E.
Henriksen
S.A.
Lundgren
B.
(
1997
)
Pneumocystis carinii from pigs and humans are antigenically distinct
.
J. Med. Vet. Mycol.
 
34
,
431
433
.
[4]
Furuta
T.
Ueda
K.
(
1987
)
Intra- and interspecies transmission and antigenic difference of Pneumocystis carinii derived from rat and mouse
.
Jpn. J. Exp. Med.
 
57
,
11
17
.
[5]
Gigliotti
F.
(
1992
)
Host species-specific antigenic variation of a mannosylated surface glycoprotein of Pneumocystis carinii
.
J. Infect. Dis.
 
165
,
329
336
.
[6]
Linke
M.J.
Cushion
M.T.
Walzer
P.D.
(
1989
)
Properties of the major antigens of rat and human Pneumocystis carinii
.
Infect. Immun.
 
57
,
1547
1555
.
[7]
Banerji
S.
Lugli
E.B.
Miller
R.F.
Wakefield
A.E.
(
1995
)
Analysis of genetic diversity at the arom locus in isolates of Pneumocystis carinii
.
J. Eukaryot. Microbiol.
 
42
,
675
679
.
[8]
Mazars
E.
Ödberg-Ferragut
C.
Dei-Cas
E.
Fourmaux
M.N.
Aliouat
E.M.
Brun-Pascaud
M.
Mougeot
G.
Camus
D.
(
1995
)
Polymorphism of Pneumocystis carinii from different host species
.
J. Eukaryot. Microbiol.
 
42
,
26
32
.
[9]
Sinclair
K.
Wakefield
A.E.
Banerji
S.
Hopkin
J.M.
(
1991
)
Pneumocystis carinii organisms derived from rat and human hosts are genetically distinct
.
Mol. Biochem. Parasitol.
 
45
,
183
184
.
[10]
Stringer
J.R.
Stringer
S.L.
Zhang
J.
(
1993
)
Molecular genetic distinction of Pneumocystis carinii from rats and humans
.
J. Eukaryot. Microbiol.
 
40
,
733
741
.
[11]
Wakefield
A.E.
Peters
S.E.
Banerji
S.
Bridge
P.D.
Hall
G.S.
Hawksworth
D.L.
Guiver
L.A.
Allen
A.G.
Hopkin
J.M.
(
1992
)
Pneumocystis carinii shows DNA homology with the Ustomycetous red yeast fungi
.
Mol. Microbiol.
 
6
,
1903
1911
.
[12]
Weinberg
G.A.
Bartlett
M.S.
(
1991
)
Comparison of pulsed field gel electrophoresis karyotypes of Pneumocystis carinii derived from rat lung, cell culture and ferret lung
.
J. Protozool.
 
38
,
64
65
.
[13]
Dei-Cas
E.
Mazars
E.
Ödberg-Ferragut
C.
Durand
I.
Aliouat
E.M.
Dridba
M.
Palluault
F.
Cailliez
J.C.
Séguy
N.
Tibayrenc
M.
Mullet
C.
Creusy
C.
Camus
D.
(
1994
)
Ultrastructural, genomic, isoenzymatic and biological features make it possible to distinguish rabbit Pneumocystis from other mammal Pneumocystis strains
.
J. Eukaryot. Microbiol.
  (
Suppl.
)
41
,
84
.
[14]
Mazars
E.
Guyot
K.
Durand
I.
Dei-Cas
E.
Boucher
S.
Ben Abderrazak
S.
Banuls
A.L.
Tibayrenc
M.
Camus
D.
(
1997
)
Isoenzyme diversity in Pneumocystis carinii from rats, mice and rabbits
.
J. Infect. Dis.
 
175
,
655
660
.
[15]
Mazars
E.
Ödberg-Ferragut
C.
Durand
I.
Tibayrenc
M.
Dei-Cas
E.
Camus
D.
(
1994
)
Genomic and isoenzymatic markers of Pneumocystis from different host species
.
J. Eukaryot. Microbiol.
  (
Suppl.
)
41
,
104
.
[16]
Lane
B.R.
Ast
J.C.
Hossler
P.A.
Mindell
D.P.
Bartlett
M.S.
Smith
J.W.
Meshnick
S.R.
(
1997
)
Dihydropteroate synthase polymorphism in Pneumocystis carinii
.
J. Infect. Dis.
 
175
,
482
485
.
[17]
The Pneumocystis International Workshop
(
1994
)
Revised nomenclature for Pneumocystis carinii
.
J. Eukaryot. Microbiol.
  (
Suppl.
)
41
,
121
122
.
[18]
Aliouat
A.
Dei-Cas
E.
Ouaissi
M.A.
Palluault
F.
Soulez
B.
Camus
D.
(
1993
)
In vitro attachment of Pneumocystis carinii from mouse and rat origin
.
Biol. Cell
 
77
,
209
217
.
[19]
Dei-Cas
E.
Mazars
E.
Aliouat
E.M.
Ödberg-Ferragut
C.
Durand
I.
Denis
C.M.
Camus
D.
(
1996
)
Infection sources, reservoir and transmission of pneumocystosis
. In:
Parasitology for the 21st Century: ICOPA VIII 1996
  (
Ozcel
Alkan
, Eds.), pp.
175
186
.
CAB International
,
Oxford
.
[20]
Peters
S.E.
Wakefield
A.E.
Whitwell
K.E.
Hopkin
J.M.
(
1994
)
Pneumocystis carinii pneumonia in thoroughbred foals: identification of a genetically distinct organism by DNA amplification
.
J. Clin. Microbiol.
 
32
,
213
216
.
[21]
Hughes
W.T.
(
1987
)
Pneumocystis carinii Pneumonitis
 .
CRC Press
,
Boca Raton, FL
.
[22]
Laakkonen
J.
Sukura
A.
(
1997
)
Pneumocystis carinii of the common shrew, Sorex araneus, shows a discrete phenotype
.
J. Eukaryot. Microbiol.
 
44
,
117
121
.
[23]
Peters
S.E.
English
K.
Laakkonen
J.
Gurnell
J.
(
1994
)
DNA analysis of Pneumocystis carinii infecting Finnish and English shrews
.
J. Eukaryot. Microbiol.
  (
Suppl.
),
41
,
108
.
[24]
Settnes
O.P.
Elvestad
K.
Clausen
B.
(
1986
)
Pneumocystis carinii Delanöe and Delanöe, 1912 found in lungs of freeliving animals in Denmark at autopsy
.
Nord. Vet. Med.
 
38
,
11
15
.
[25]
Mazars
E.
Guyot
K.
Fourmaintraux
S.
Renaud
F.
Pétavy
F.
Camus
D.
Dei-Cas
E.
(
1997
)
Detection of Pneumocystis in European wild animals
.
J. Eukaryot. Microbiol.
 
44
(
Suppl
),
39
.
[26]
Aliouat
E.M.
Mazars
E.
Dei-Cas
E.
Cesbron
J.Y.
Camus
D.
(
1993
)
Intranasal inoculation of mouse, rat or rabbit-derived Pneumocystis in SCID mice
.
J. Protozool. Res.
 
3
,
94
98
.
[27]
Aliouat
E.M.
Mazars
E.
Dei-Cas
E.
Delcourt
P.
Billaut
P.
Camus
D.
(
1994
)
Pneumocystis cross infection experiments using SCID mice and Nude rats as recipient host, showed strong host-species specificity
.
J. Eukaryot. Microbiol.
  (
Suppl.
)
41
,
71
.
[28]
Furuta
T.
Fujita
M.
Mukai
R.
Sakakibara
I.
Sata
T.
Miki
K.
Hayami
M.
Kojima
S.
Yoshikawa
Y.
(
1993
)
Severe pulmonary pneumocystosis in simian acquired immunodeficiency syndrome induced by simian immunodeficiency virus — its characterization by the polymerase-chain-reaction method and failure of experimental transmission to immunodeficient animals
.
Parasitol. Res.
 
79
,
624
628
.
[29]
Gigliotti
F.
Harmsen
A.G.
Haidaris
C.G.
Haidaris
P.J.
(
1993
)
Pneumocystis carinii is not universally transmissible between mammalian species
.
Infect. Immun.
 
61
,
2886
2890
.
[30]
Mazars
E.
Herbecq
S.
Szypura
A.S.
Fruit
J.
Camus
D.
Dei-Cas
E.
(
1996
)
Pneumocystis carinii detection in 158 HIV-seronegative patients
.
J. Eukaryot. Microbiol.
  (
Suppl.
)
43
,
28
.
[31]
Wakefield
A.E.
(
1996
)
DNA sequences identical to Pneumocystis sp.f. carinii and Pneumocystis carinii sp.f. hominis in samples of air spora
.
J. Clin. Microbiol.
 
34
,
1754
1759
.
[32]
Dei-Cas
E.
Vernes
A.
(
1986
)
Parasitic adaptation of pathogenic fungi to the mammalian hosts
.
CRC Crit. Rev. Microbiol.
 
13
,
173
218
.
[33]
Dei-Cas
E.
Mazars
E.
Aliouat
E.M.
Nevez
G.
Cailliez
J.C.
Camus
D.
(
1998
)
The host-specificity of Pneumocystis carinii
.
J. Mycol. Méd.
 
8
,
1
6
.
[34]
Soulez
B.
Palluault
F.
Cesbron
J.Y.
Dei-Cas
E.
Capron
A.
Camus
D.
(
1991
)
Introduction of Pneumocystis carinii in a colony of SCID mice
.
J. Protozool.
  (
Suppl.
)
38
,
123
125
.
[35]
Ceré
N.
Drouet-Viard
F.
Dei-Cas
E.
Chanteloup
N.
Coudert
P.
(
1997
)
In utero transmission of Pneumocystis carinii sp.f. oryctolagi
.
Parasite
 
4
,
325
330
.
[36]
Tsolaki
A.G.
Miller
R.F.
Underwood
A.P.
Banerji
S.
Wakefield
A.E.
(
1996
)
Genetic diversity at the internal transcribed spacer regions of the ribosomal RNA operon among isolates of Pneumocystis carinii from AIDS patients with recurrent pneumonia
.
J. Infect. Dis.
 
174
,
141
156
.
[37]
Nevez
G.
Jounieaux
V.
Linas
M.D.
Guyot
K.
Léophonte
P.
Massip
P.
Schmit
J.L.
Séguéla
J.P.
Camus
D.
Dei-Cas
E.
Raccurt
C.
Mazars
E.
(
1997
)
High frequency of Pneumocystis carinii sp. f. carinii colonization in HIV-negative patients
.
J. Eukaryot. Microbiol.
  (
Suppl.
)
44
,
36
.
[38]
Stringer
J.R.
Wakefield
A.E.
Cushion
M.
Dei-Cas
E.
(
1997
)
Pneumocystis taxonomy and nomenclature: an update
.
J. Eukaryot. Microbiol.
  (
Suppl.
)
44
,
5
6
.
[39]
Mayr
E.
(
1942
)
Systematics and the Origin of Species
 .
Columbia University Press
,
New York
.
[40]
Wakefield
A.E.
Stringer
J.R.
Tamburrini
E.
Dei-Cas
E.
(
1998
)
Genetics, metabolism and host specificity of Pneumocystis carinii
.
J. Med. Vet. Mycol.
  (
in press
).
[41]
Cailliez
J.C.
Séguy
N.
Denis
C.M.
Aliouat
E.M.
Mazars
E.
Polonelli
L.
Camus
D.
Dei-Cas
E.
(
1996
)
Pneumocystis carinii: an atypical fungal microorganism
.
J. Med. Vet. Mycol.
 
34
,
227
239
.
[42]
Stringer
J.R.
Walzer
P.D.
(
1996
)
Molecular biology and epidemiology of Pneumocystis carinii infection in AIDS
.
AIDS
 
10
,
561
571
.
[43]
Stringer
J.R.
(
1996
)
Pneumocystis carinii: what is it, exactly?
Clin. Microbiol. Rev.
 
9
,
489
498
.