American Journal of Plant Sciences, 2011, 2, 416-424
doi:10.4236/ajps.2011.23047 Published Online September 2011 (http://www.SciRP.org/journal/ajps)
Analysis of Genetic Diversity in Poeciloneuron
pauciflorum Bedd.―An Endemic Tree Species
from the Western Ghats of India
Padmesh Pandaram Pillai1*, Jayakumari Sanakan Sajan1, Kuttapetty Manikantan Menon1,
Achuthan Sudarshanan Hemanthakumar1, Alagramam Govindasamy Pandurangan1,
Peringatulli Narayanan Krishnan1, Soorimuthu Seeni2
1Tropical Botanic Garden and Research Institute, Thiruvananthapuram, India; 2Sathyabama University, Chennai, India.
Received April 18th, 2011; revised May 6th, 2011; accepted May 26th, 2011.
ABSTRACT
Poeciloneuron pauciflorum is a narrow endemic having highly restricted distribution in the southern-western Ghats
region of India. 18 accessions of P. pauciflorum collected from four different locations were analyzed for genetic varia-
tion using 20 random primers. Out of 14 1 amplicons generated, 130 of them were polymorphic (92.20% polymorphism).
Contrary to the general concept of low genetic variation associated with endemic plant species, P. pauciflorum exhibits
high genetic diversity as the similarity index value based on Nei & Li’s similarity coefficient ranges from 0.36 to 0.95
with mean GS = 0.72 and Shannon’s information measure (0.43). Cluster analysis showed grouping of all accessions
from the State of Tamil Nadu into two major clusters with few outliers while those from the State of Kerala also clus-
tered with them. Accessions from Kalliankadu forest segment harbors maximum diversity as indicated by various ge-
netic diversity indices like h, I, Ht, Hs, Gst, PL and hence this site is recommended for in situ conservation of this nar-
row endemic. The main factors responsible for the high level of genetic diversity among accessions are probably re-
lated to the reproductive isolation and ecological breadth. The strong genetic variability among accessions indicates
that the management for the conservation of the genetic diversity in P. pauciflorum should aim to preserve every acces-
sion. The present study assumes significance as it negotiates endemism and genetic variation in tree species, a global
phenomenon having wide implications in species diversity and conservation.
Keywords: RAPD, Neoendemism, Xanthones
1. Introduction
Poeciloneuron pauciflorum Bedd.―(Clusiaceae) is a
critically endangered tree species endemic to India hav-
ing sparse distribution in the Southern Western Ghats of
Kerala and Tamil Nadu States [1]. It is a medium sized
tree growing up to 15 m tall and 2 m in girth, distributed
in the evergreen forest on the hills, particularly along
riverbanks or watercourse between 600 to 1500 MSL al-
titude. The species is seen in KMTR (Kalakadu Mudan-
thurai Tiger Reserve) forest segment of Tamil Nadu. Re-
cently 12 new additional sites grouped under three major
regions viz. Mahendragiri, Chambakuchi and Kallar
comprising the Kanyakumari Wildlife Sanctuary of Ta-
mil Nadu [2] and from Chemungi-Attayar forest region of
Kerala [3] has been reported. The species yields economi-
cally important timber with the wood reddish in
colour, hard and heavy. The genus Poeciloneuron with
the species P. indicum was initially considered as mono-
typic by Beddome [4] under the family Tenstromeiaceae
and later in 1871 added P. pauciflorum as the second
species. However, the genus with just two species finds
itself in taxonomic reassessments based on morphologi-
cal, wood anatomical and palynological data [5] as the
extent of variation at the interspecific level is estimated
to be high.
Like few other endemics of the Western Ghats, P.
pauciflorum shows large amount of variation in plant
morphology and is threatened by habitat loss. The family
Clusiaceae is known for high amount of xanthones [6,7]
and P. pauciflorum is no exception as it is a rich source
of xanthones like (1,6-dihydroxy-7-methoxyxanthone and
1,6-dihydroxy-7-methoxyxanthone 6-O-β-d-glucoside) in
addition to 12 others, such as 1,5-dihydroxy-, 1,5-dihy-

Analysis of Genetic Diversity in Poeciloneuron Pauciflorum Bedd. 417
―An Endemic Tree Species from the Western Ghats of India
droxy-3-methoxy-, 1,7-dihydroxy-, 1-hydroxy-7-methoxy-,
2-methoxy-, 4-methoxy-, 1,4,5-trihydroxy-, 1,3,5-trihy-
droxy-, 1,3,6-trihydroxy-7-methoxy-, 1,3,7-trihydroxy-, 3-
hydroxy-2-methoxyxanthone and (−) -epicatechin [8].
These xanthones show various bioactivities like an-
timalarial [9], effective against anti-methicillin-resistant
Staphylococcus aureus [10-12], tumor-promoting inhibi-
tion [13], selective cyclooxygenase-2 inhibition [14] and
inhibitory effects on PAF-induced hypotension [15]. Be-
sides, they are used for the treatment of mental disorder,
infectious diseases and for exorcism activities by the
Kani tribals of South India [16]. According to IUCN
conservation assessment the plant species is red listed as
endemic and critically endangered (Poeciloneuron pau-
ciflorum. In: IUCN 2010. IUCN Red List of Threatened
Species. Version 2010.4. <www.iucnredlist.org> as on
09 May 2011). Few fragmented populations distributed
in a small geographical region entails an urgent reas-
sessment of the threat category of this species for which
we have employed RAPD as tool for estimation of ge-
netic variation in samples collected from few locations
across the region. RAPD as a cost effective DNA marker
system has been successfully employed to detect varia-
tion at the inter-specific [17-19], intraspecific [20], in-
tervarietal [21], intergeneric genome analysis [22] and at
the cultivar levels [23].
The present study assumes significance as it negotiates
endemism and genetic variation in tree species, a global
phenomenon having wide implications in species diver-
sity and conservation.
2. Materials and Methods
2.1. Collection of Plant Materials
Total of 18 accessions of Poeciloneuron pauciflorum were
collected from parts of Tamil Nadu and Kerala. As the
tree species is highly endemic, the distance between col-
lection sites (inter-population) is approximately 20 km in
Tamil Nadu and within each site (intra-population) ac-
cessions were collected at a distance of 0.2 km to 0.5 km.
The distance between Kerala and Tamil Nadu forest
segment is about 20 km. Populations of P. pauciflorum
were located in four locations of Tamil Nadu and Kerala
viz. Inchikuzhi, Veliyar and Kalliankadu in Kalakkadu-
Mundanthurai Tiger Reserve (Tamil Nadu) and Attayar
in Peppara Wild Life Sanctuary (Kerala) (Figure 1).
Figure 1. Map showing the Western Ghats and the distribution of Poeciloneuron pauciflorum (insight).

Analysis of Genetic Diversity in Poeciloneuron Pauciflorum Bedd.
418
―An Endemic Tree Species from the Western Ghats of India
2.2. Genomic DNA Isolation and RAPD
Total genomic DNA from the young leaves was isolated
following the modified Murray and Thompson [24] me-
thod using CTAB. After ethanol precipitation DNA was
resuspended in 100 µl of 1xTE buffer (pH 8.0). The
DNA was quantified spectrophotometrically by taking
the absorbance at 260 nm using Biophotometer (Ep-
pendorf, Hamburg). RAPD assay was carried out in 25 µl
reaction mixture containing 0.5 µl dNTPs (0.2 mM), 2.5 µl
10X polymerase buffer (10 mM Tris-HCl, 1.5 mM
MgCl2, 50 mM KCl, 0.1% Triton X-100), 0.5 µl Taq
DNA po- lymerase (1.0 U/µl) (Finnzymes, Helsinki,
Finland), 1 µl primers(15 pmol) from Kit “C”, Kit “D”
and Kit “S” (Biogene, USA) and 50 ng of genomic DNA.
The final volume was made up to 25 µl by adding milli Q
water. The amplification was performed in a thermal
cycler (Eppendorf-ESP-S, Hamburg) with oil free opera-
tion. After the initial cycle of 2 min at 94˚C, 2 min at
36˚C and 2 min at 72˚C, 38 cycles of 1 min at 94˚C, 1
min at 36˚C and 2 min at 72˚C were performed. The last
cycle was followed by 7 min extension at 72˚C. Reaction
mix- ture wherein template DNA replaced by distilled
water was used as negative control. Amplified products
were resolved in 1.4% agarose gel (1x TBE) followed by
EtBr staining.
2.3. Genetic Data Analysis
Amplification with each random primer (45 primers) was
repeated 3 times and those primers that produced repro-
ducible and consistent bands (20 primers) were selected
for data generation. Reproducible RAPD products were
scored against the presence or absence of a fragment and
denoted as “+” or “–”, respectively. Dice coefficient of
similarity defined as 2a/(2a + u), where “a” is the number
of positive matches and “u” is the number of non
matches was computed using the WINDIST software.
The scored binary matrix was analyzed for the construc-
tion of phenogram and determination of confidence lim-
its by bootstrap analysis using the WINBOOT software
[26]. The genetic variation was analyzed for various pa-
rameters. The genotype and allelic frequency data were
used to compute the genetic diversity indices i.e. ob-
served number of alleles (na), expected number of alleles
(ne), Shannon index of genetic diversity (I), nei’s gene
diversity (h) at the population level, (Ht) heterozygosity
at the polymorphic loci, average heterozygosity (Hs) and
degree of genetic differentiation (Gst), using the statisti-
cal package POPGENE 1.3 [27]. The populations from
which the samples taken for the present analysis were
assumed to be in Hardy-Weinberg equilibrium implying
that the population is at random mating. Based on the
above assumption, the bands were scored and estimation
of heterozygosity (Ht) was done according to the formula:
Ht = 1 – ∑pi
-2 where pi is the frequency of the ith allele in
the population.
3. Results
3.1. RAPD Polymorphism
The 20 random primers used for the estimation of intras-
pecific variation in 18 samples of P. pauciflorum (Tables
1 and 2) provide interesting insights into the prevailing
Table 1. List of Poeciloneuron pauciflorum samples with
details of collection site and location coordinate.
Sl No.Sample ID Location Latitude/Longitude
1 Pp 1 Near Veliyar
8˚36.651N
77˚15.564E
2 Pp 2 Near Veliyar 8˚36.682N
77˚15.531E
3 Pp 3 Near Veliyar 8˚36.642N
77˚15.574E
4 Pp 4 Near Veliyar 8˚36.694N
77˚15.521E
5 Pp 5 Near Veliyar 8˚36.738N
77˚15.794E
6 Pp 6 Near Veliyar 8˚36.721N
77˚15.564E
7 Pp 7 Near Veliyar 8˚36.788N
77˚15.720E
8 Pp 8 Kalliankadu 8˚37.482N
77˚16.432E
9 Pp 9 Kalliankadu 8˚37.456N
77˚16.414E
10 Pp 10 Kalliankadu 8˚37.489N
77˚16.112E
11 Pp 11 Kalliankadu 8˚37.542N
77˚16.342E
12 Pp 12 Inchikuzhi 8˚37.484N
77˚18.094E
13 Pp 13 Inchikuzhi 8˚37.454N
77˚18.014E
14 Pp 14 Inchikuzhi 8˚37.521N
77˚18.014E
15 Pp 15 Inchikuzhi 8˚37.621N
77˚18.421E
16 Pp 16 Inchikuzhi 8˚37.458N
77˚18.261E
17 Pp 17 Attayar 8˚41.117N
77˚11.369E
18 Pp 18 Attayar 8˚41.186N
77˚11.314E

Analysis of Genetic Diversity in Poeciloneuron Pauciflorum Bedd. 419
―An Endemic Tree Species from the Western Ghats of India
Table 2. List of primers and their sequence used for RAPD
analysis of Poeciloneuron pauciflorum.
Sl No. Primers Primer
sequence
5'3'
No. of
bands
No. of
polymorphic
bands
1 S61 TTCGAGCCAG 11 11
2 S62 GTGAGGCGTC 6 4
3 S63 GGGGGTCTTT 9 9
4 S64 CCGCATCTAC 7 7
5 S65 GATGACCGCC 5 3
6 S66 GAACGGACTC 9 8
7 S68 TGGACCGGTG 5 5
8 S69 CTCACCGGTG 8 6
9 S70 TGTCTGGGTG 6 6
10 S71 AAAGCTGCGG 9 7
11 C73 AAGCCTCGTC 8 8
12 C74 TGCGTGCTTG 9 8
13 C75 GACGGATCAG 9 9
14 C76 CACACTCCAG 6 5
15 S-80 ACTTCGCCAC 6 6
16 D-08 GTGTGCCCCA 6 6
17 D-10 GGTCTACACC 5 5
18 D-13 GGGGTGACGA 5 5
19 D-17 TTTCCCACGG 6 6
20 D-18 GAGAGCCAAC 6 6
Total No. of bands 141 130
Mean per primer 7.05 6.50
genetic variability in this narrow endemic tree. Out of 141
products generated, 130 were polymorphic (92.20%). On
an average, the primers generated 7.05 amplicons and
6.50 polymorphism per primer. The number of products
generated by these arbitrary 10-mer primers was found to
range from 5 to 11 with primer S61 giving the maximum
(11) and primer S65, D10 and D13 giving the minimum
(5) number of amplicons. While majority of the primers
produced 100% polymorphism, there was not a single
primer that resulted in complete monomorphism. The
similarity matrix developed using the WINDIST soft-
ware showed similarity index ranging from 0.36 to 0.95
with mean value of 0.72 (Table 3). Nei’s gene diversity
at population level (h), Shannon index (I), expected
number of alleles (ne) were calculated to estimate genetic
variation. The accessions included in this study showed
relatively high level of genetic diversity, i.e. h = 0.3 and I
= 0.43. The mean genetic diversity based on Nei’s statis-
tics [28] also supports the other data. The mean value of
heterozygosity (Ht) observed in the various accessions of
P. pauciflorum was found to be 0.45. The mean value of
average heterozygosity value was 0.30. The heterozygos-
ity values and degree of genetic differentiation (Gst) is
shown in Tables 4(a)-(b). The other diversity measures
as indicated in Tables 4(a)-(b) also revealed more diver-
sity at the inter and intra population levels. The gene
flow (Nm) among all accessions is 0.98, calculated on
the assumption that the accession under study follows the
inland model [29] which predicts a simple relationship
between the numbers of migrants an accession receives
per generation.
Fst = 1/4 (Nm+1) from which Nm was derived as Nm
= (1 – Fst)/4Fst. The Gst value obtained from the POP-
GENE analysis was substituted for Fst and derived the
rate of gene flow [30].
Table 3. Similarity matrix of accessions of Poeciloneuron pauciflorum analyzed using dice’s coefficient.
1.00
0.91 1.00
0.89 0.94 1.00
0.87 0.92 0.92 1.00
0.91 0.93 0.96 0.90 1.00
0.90 0.89 0.93 0.89 0.95 1.00
0.80 0.81 0.83 0.78 0.81 0.77 1.00
0.92 0.86 0.85 0.83 0.88 0.88 0.76 1.00
0.55 0.55 0.53 0.58 0.54 0.55 0.54 0.55 1.00
0.83 0.79 0.80 0.79 0.82 0.81 0.71 0.85 0.55 1.00
0.82 0.83 0.80 0.77 0.82 0.79 0.78 0.84 0.56 0.82 1.00
0.70 0.70 0.69 0.66 0.71 0.71 0.61 0.72 0.40 0.69 0.72 1.00
0.68 0.68 0.68 0.65 0.70 0.68 0.63 0.70 0.38 0.67 0.72 0.94 1.00
0.62 0.63 0.63 0.58 0.64 0.62 0.60 0.62 0.36 0.62 0.63 0.80 0.79 1.00
0.65 0.68 0.68 0.65 0.70 0.69 0.62 0.68 0.41 0.67 0.67 0.90 0.88 0.84 1.00
0.53 0.56 0.58 0.56 0.59 0.55 0.46 0.56 0.46 0.55 0.57 0.72 0.71 0.61 0.75 1.00
0.65 0.65 0.68 0.63 0.68 0.66 0.60 0.63 0.44 0.63 0.62 0.76 0.75 0.65 0.72 0.78 1.00
0.62 0.65 0.65 0.63 0.65 0.63 0.58 0.59 0.45 0.63 0.61 0.72 0.70 0.67 0.70 0.71 0.85 1.00
P1 P2 P3 P4 P5 P6 P7 P8 P9 P10 P11 P12 P13 P14 P15 P16 P17 P18

Analysis of Genetic Diversity in Poeciloneuron Pauciflorum Bedd.
420
―An Endemic Tree Species from the Western Ghats of India
Table 4. (a) Mean genetic Diversity of 18 accessions of Poeciloneuron pauciflorum based on Nei’s (1987) statistics; (b) Multi-
population analysis showing mean value of Genetic diversity indices.
(a)
Ht Hs Gst Nm na ne h I
Mean
0.45 0.29 0.33 0.98 1.73 1.54 0.30 0.43
(b)
Population h I Nm Gst Ht Hs PL % PL
Veliyar 0.33 0.49 3.13 0.14 0.33 0.28 18 90.00
Kalliankadu 0.43 0.61 1.23 0.29 0.43 0.30 19 95.00
Inchikuzhi 0.34 0.50 1.91 0.21 0.34 0.27 18 90.00
Attayar 0.26 0.39 2.2 0.04 0.26 0.25 15 75.00
Ht-heterozygosity at the polymorphic loci, Hs-average heterozygosity, Gst-degree of genetic differentiation, na-observed number of alleles, ne-expected number
of alleles, h-Nei’s gene diversity at population level, I-Shannon index of genetic diversity, PL-Polymorphic Loci, %PL-percentage Polymorphic Loci.
3.2. Cluster Analysis
The samples of Poeciloneuron clustered broadly under
two major groups with sub-clusters within each group.
All the samples except one from Veliyar grouped and
constituted cluster I at a moderate confidence interval
limit of 78.2% while samples from Inchikuzhi formed
cluster II along with the two samples of Attayar at 92.7%
confidence limit (Figure 2). The grouping of Inchi- kuz-
hi and Attayar samples showed high robustness as they
were supported at 92.7% and 94.6% confidence interval
limits, respectively. All the four samples from Kal-
liankadu were found to form outliers. Three samples
were placed characteristically between the two clusters
and one outside the clusters (P-9).
4. Discussion
It is well known that the nature and distribution of ge-
netic variability within and among natural populations of
species constitutes its genetic structure which in turn is
affected by edaphic/demographic factors [31,32] and
evolutionary processes [33]. The characteristic genetic
structure of a population reflects the interactions of vari-
ous factors like long-term evolutionary history of the
species (shifts in the distribution, habitat distribution,
habitat fragmentation and population isolation), genetic
drift, mating system, gene flow and selection [34]. How-
ever, attributes like the progenitor, probability of com-
mon origin, kin structure and inbreeding within popula-
tions all have significant effects on genetic differentiation
among populations [35].
Above all, the extent of geographical distribution of
the species is yet another single major factor that deter-
mines its variability [36-39]. In general, it is accepted
that long term conservation strategies for plant species
demands better understanding of the ecological and ge-
netical variables prevailing in that particular niche over a
period of time. Therefore, genetic variability analysis has
significant implication in designing conservation strate-
gies for species which are endemic and endangered in
status.
Genetic Variation in Narrow Endemics
The RAPD data generated out of 20 random primers for
P. pauc iflorum collected from forest segments that span
across two southern most States of India provided inter-
esting insights into the existing diversity in this highly
endemic tree species. The coefficient of genetic similar-
ity ranging from 0.36 to 0.95 with mean value of 0.72
suggests high variability in the species despite its en-
demic status. The grouping of samples apparently re-
flects characteristic fragmentation of the population with
all samples from Tamil Nadu side of the Western Ghats
forming two independent clusters at the two ends of the
phenogram with few samples forming outliers in between
them. Those samples from the Kerala side of the Western
Ghats though less in number tend to form grouping with
the samples from Inchikuzhi. Multi-population analysis
to determine intra and inter population diversity shows
that most of the diversity indices (Ht, Hs, Gst, Nm, h, I,
PL) are high for samples from Kaliankadu followed by
Inchikuzhi, Veliyar and Attayar. It is generally accepted
that endemics tend to have lower levels of genetic varia-
tion than their wide spread congeners [40], a condition
often attributed to reproductive isolation and geographi-
cal fragmentation. Endemics are generally considered to
be inbreeders as their progenitors as founders were as-
sumed to have benefited from self compatibility. It is

Analysis of Genetic Diversity in Poeciloneuron Pauciflorum Bedd. 421
―An Endemic Tree Species from the Western Ghats of India
presumed that at a later stage of divergence they became
reproductively isolated from the parent population and
resorted to inbreeding as the only possible mode of re-
production. There are many schools of thought that en-
dorse low levels of genetic variation in endemic plant
species based largely on geographical distribution pattern
and reproductive isolation. In contrast to our expectation
of low genetic diversity, P. pauciflorum shows high ge-
netic diversity and the mean genetic diversity values.
Nei’s statistics also support this level of genetic diversity.
High genetic differentiation (Gst = 0.33) and low gene
flow (Nm = 0.98) indicates the possible threat to the taxa
in spite of its present diverse nature showing strong ten-
dency to genetic drift and inbreeding depression unless
measures are taken for introducing sufficient number of
migrants into the population. Despite its endemic status
and restricted geographical range P. pauciflorum shows
high level of genetic diversity and the two major genetic
consequences of small population size for long periods of
time are high levels of genetic drift and inbreeding [41,
42]. Inbreeding species maintain relatively more of their
genetic diversity among populations rather than within
populations than do out crossers [43]. Some of the fac-
tors such as recent speciation from a more wide-spread
species, recent changes in distribution or habitat, breed-
ing system, somatic mutations, multiple founder events,
tropical forest fragmentation leading to decreased gene
flow, increased inbreeding producing a high differentia-
tion among remnant populations, invoked to explain high
genetic diversity of the narrow endemic species like,
Delpbinuim viridescens [44], Symplocos laurina, Eurya
nitada [45], Primula interjacens [46], Abeliophyllum dis-
tichum [47], and Antirhea aromatica [48] may also ex-
plain the observed endemism of P. pauciflorum.
Figure 2. Phenogram based on UPGMA analysis of Poeciloneuron pauciflorum accessions. Numbers at the fork indicate boot-
strap values.

Analysis of Genetic Diversity in Poeciloneuron Pauciflorum Bedd.
422
―An Endemic Tree Species from the Western Ghats of India
Accordingly, narrow endemics are mostly referred as
products of speciation that differ in their origin and status
of rarity only in certain aspects that in turn are controlled
by such factors as geographic area, ecological breadth,
reproductive isolation and most importantly the amount
of genetic variation inherent in the taxa [49]. In fact, the
first three factors defines the genetic structure of the taxa
to a considerable extent that perpetuation and establish-
ment relies mostly on the extent of genetic variation.
The niche variation hypothesis which refers to species
adaptation to narrow ecological conditions also explains
the significant role of heterozygosity in genetic diversity
and endemism and thereby the genesis of narrow endem-
ics. Thereby other major factor besides selection pressure
that determine heterozygosity is effective population size
which in turn could be the product of (a) smaller total
population size compared to those species having cos-
mopolitan distribution (b) genetic bottle neck at the time
of origin of the species which in due course with rapid
speciation may produce a certain low level of genetic
variation, a condition that could be transitory (c) in-
breeding/selfing may reduce effective population size
and hence heterozygosity and (d) population fragmenta-
tion due to geographical or reproductive isolation. We
assume that some or all of these factors must have con-
tributed to the peculiar distribution and current status of
endemism seen in P. pauciflorum. The populations from
which the samples taken for the present analysis were
assumed to be in Hardy-Weinberg equilibrium implying
that the population is at random mating. Though direct
measure of heterozygosity was not calculated in the pre-
sent study the indirect measures reflected through various
diversity indices clearly demonstrates high variability in
the taxa that is not otherwise expected in this category of
narrow endemics in general and in a taxon like P. pau-
ciflorum in particular where the natural mode of repro-
duction is through selfing as indicated by the floral
structure (bi-sexual flowers, stigma acute wherein both
stigma and anther are placed in the same level suggestive
of inbreeding). Therefore the general concept of associ-
ating low genetic variation in natural populations of plant
species with endemism entails further extrapolation to
negotiate the prevalence of unseemingly high diversity in
this species. However such situations of associating en-
demism with high genetic diversity has been earlier re-
ported in vascular plants like rare ferns Adenophorus
periens [50], endemic Agave victoriae-reginae [51], nar-
row endemic tree species Antirhea aromatica, Antir-
rhinum charidemi and A. valentinum [52], narrow en-
demic conifer Cupressus macrocarpa, which has highly
restricted distribution shows fairly good variability. All
seven species of the Californian endemic grass genus
Orcuttia that have as much genetic variability as any
other widely distributed members of Graminae [53].
Among these, Antirhea aromatica has exceptionally high
genetic diversity and variability, despite its low popula-
tion density [48]. The phenomenon is likely to be associ-
ated with the reproductive system as in certain members
of the family Rubiaceae, there is pre-zygotic incompati-
bility crossing system which reduces inbreeding and
thereby loss of genetic diversity. We also propose similar
or associated mechanism underlying the reproductive
biology of P. pauciflorum that reduces the possibility of
selfing and maintains the observed levels of genetic di-
versity in this narrow endemic species.
In conclusion our results show that this narrow en-
demic tree species has generated a systematic process of
genetic isolation yet capable of maintaining genetically
viable populations in few patches of the Western Ghats
region. Further the data is suggestive that in situ con-
servation may be adopted for those populations in Kal-
liankadu as they are shown to harbor maximum amount
of variability possibly present in the species. We also re-
commend an ex situ conservation strategy through con-
ventional or non-conventional approach involving simple
micropropagation to increase the number of plantlets and
thereby delist the species from its current endemic status.
6. Acknowledgements
The authors greatly acknowledge the Kerala and Tamil
Nadu Forest Departments for granting permission to col-
lect plant samples from the respective forest segments.
Also, PP, AGP, PNK and SS thank the Department of
Biotechnology, Government of India for the research
grant (Grant No. BT/PR7058/BCE/08/440/2006).
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Abbreviations: RAPD: Random Amplified Polymorphic DNA; GS: Genetic Similarity Coefficient.