Indian J Exp Biol
JULY 2002
CODEN: IJEB (A6) 40(7) 749-860 (2002)
ISSN: 0019-5189
Indian Journal of Experimental Biology
http : // www.niscom.res.in; http : // www.bioline.org.br/
VOLUME 40
NUMBER 7
CODEN : IJEB (A6) 40(7) 749-860 (2002)
JULY 2002
ISSN : 0019-5189
CONTENTS
View Point
Novel parasite (nematode) transglutaminase  Potential drug target
753
R Chandrashekar
Review Articles
Role of rhizobial biosynthetic pathways of amino acids, nucleotide bases and vitamins in symbiosis
Gursharn S Randhawa & Raad Hassani
Validation of traditional claim of Tulsi, Ocimum sanctum Linn. as a medicinal plant
S K Gupta, Jai Prakash & Sushma Srivastava
755
765
Papers
Localisation of identical organophosphorus pesticide degrading (opd) genes on genetically dissimilar
indigenous plasmids of soil bacteria: PCR amplification, cloning and sequencing of opd gene
from Flavobacterium balustinum
Sita Somara, Bramanandam Manavathi, Christoph C Tebbe &
Dayananda Siddavatam
774
Short-term androgen deprivation does not alter CaR and VDR mRNA expression in duodenal
mucosa in male rats
S Tiwari, S K Gupta, M Mehrotra, G Agarwal , P K Awasthi & M M Godbole
780
Effect of phosphocreatine on H+ extrusion, pHi and dimorphism in Candida albicans
Nikhat Manzoor, M Amin & Luqman A Khan
Control of hyperglycemia and retardation of cataract by mulberry (Morus indica L.) leaves in
streptozotocin diabetic rats
B Andallu & N Ch Varadacharyulu
785
791
Growth behaviour and bioproduction of indole acetic acid by a Rhizobium sp. isolated from root
nodules of a leguminous tree Dalbergia lanceolaria
A C Ghosh & P S Basu
796
Secretion of ligninperoxidase by Penicillium citrinum, Fusarium oxysporum and
Aspergillus terreus
Meera Kumari, R S S Yadav & K D S Yadav
802
(Contd)
Effect of low level exposure of lead and cadmium on hepatic estradiol metabolism in female rats
Anil Pillai, Laxmipriya, Ami Rawal & Sarita Gupta
Screening of Indian plants for biological activity: Part XVI
A K Goel, D K Kulshreshtha, M P Dubey & S M Rajendran
807
812
Notes
Antimicrobial potentiality of a phenothiazine group of antipsychotic drug-prochlorperazine
Rupa Mazumder, Sharmila Ray Chaudhuri & Avijit Mazumder
Synergistic effect of ayurvedic pearl preparation on enhancing effectiveness of
antibiotics
Manisha Kulkarni, J Y Deopujari & H J Purohit
828
831
SEM study on cytotoxic effect of monocrotophos (MCP) on lungs of rat
Sangeeta, S M Handa & P K Mittal
835
Use of RAPD in assessing the genetic variability in Spodoptera litura
S Janarthanan, S Seshadri, K Kathiravan & S Ignacimuthu
839
Entomopoxvirus of cotton bollworm, Helicoverpa armigera (Hbn.)
K Narayanan
842
Mass production of polyhedral occlusion bodies of NPV of Helicoverpa armigera in relation to
dose, age and larval weight
K Narayanan & S Jayaraj
846
In vitro regeneration of Acacia catechu Willd. from callus and mature nodal
explants  An improved method
Manisha Thakur, D R Sharma, Kamlesh Kanwar & Anil Kant
850
Cyanobacterial N2 fixation in presence of nitrogen fertilizers
A E Mekonnen, Radha Prasanna & B D Kaushik
854
Effect of lead on Na+, K+-ATPase activity in Penaeus indicus postlarvae
C Satyavathi & Y Prabhakara Rao
858
Indian Journal of Experimental Biology
Vol. 40, July 2002, pp. 753-754
View Point
Novel parasite (nematode) transglutaminase  Potential drug target
R Chandrashekar
Indian Journal of Experimental Biology
Vol. 40, July 2002, pp. 755-764
Review Articles
Role of rhizobial biosynthetic pathways of amino acids, nucleotide bases
and vitamins in symbiosis
Gursharn S Randhawa* & Raad Hassani§
Rhizobia require the availability of 20 amino acids for the establishment of effective symbiosis with legumes. Some of
these amino acids are synthesized by rhizobium, whereas the remaining are supplied by the host plant. The supply from
plant appears to be plant-type specific. Alfalfa provides arginine, cysteine, isoleucine, valine and tryptophan, and cowpea
and soybean provide histidine. The production of ornithine and anthranilic acid, the intermediates in the biosynthetic
pathways of arginine and tryptophan, respectively, seems to be essential for effective symbiosis of Sinorhizobium meliloti
with alfalfa. The expression of ilvC gene of S. meliloti is required for induction of nodules on the roots of alfalfa plants. An
undiminished metabolic flow through the rhizobial pathways for the synthesis of purines and pyrimidines and the synthesis
of biotin, nicotinic acid, riboflavin and thiamine by rhizobium appear to be requirements for normal symbiosis. To the best
of our knowledge, this is the first review article on the role of rhizobial biosynthetic pathways of amino acids, nucleotide
bases and vitamins in rhizobium-legume symbiosis. The scientific developments of about 35 years in this field have been
reviewed.
Indian Journal of Experimental Biology
Vol. 40, July 2002, pp. 765-773
Validation of traditional claim of Tulsi, Ocimum sanctum Linn.
as a medicinal plant
S K Gupta*, Jai Prakash & Sushma Srivastava
In several ancient systems of medicine including Ayurveda, Greek, Roman, Siddha and Unani, Ocimum sanctum has
vast number of therapeutic applications such as in cardiopathy, haemopathy, leucoderma, asthma, bronchitis, catarrhal fever,
otalgia, hepatopathy, vomiting, lumbago, hiccups, ophthalmia, gastropathy, genitourinary disorders, ringworm, verminosis
and skin diseases etc. The present review incorporates the description of O. sanctum plant, its chemical constituents, and
various pharmacological activities.
Papers
Indian Journal of Experimental Biology
Vol. 40, July 2002, pp. 774-779
Localisation of identical organophosphorus pesticide degrading (opd) genes on
genetically dissimilar indigenous plasmids of soil bacteria: PCR amplification,
cloning and sequencing of opd gene from Flavobacterium balustinum
Sita Somara, Bramanandam Manavathi, Christoph C Tebbe & Dayananda Siddavatam
Received 19 November 2001; revised 28 February 2002
Plasmid borne organophosphorus pesticide degrading (opd) gene of Flavobacterium balustinum has been amplified
using polymerase chain reaction (PCR) and the resulting PCR product (1.25 Kb) was cloned in pUC18. Further, a detailed
restriction map was determined to PCR product and subcloned as overlapping restriction fragments. The nucleotide
sequence was determined for all subclones to obtain complete sequence of PCR amplified fragment. The sequence showed
98% similarity to opd genes cloned from other soil bacteria isolated from diversified geographical regions. The protein
sequence predicted from the nucleotide sequence was almost identical to parathion hydrolase, a triesterase involved in
hydrolysis of triester bond found in variety of op-pesticides. The signal sequence of parathion hydrolase contained recently
discovered twin arginine transport (tat) motif. It appears that tat motif plays a critical role in membrane targeting of
parathion hydrolase.
Indian Journal of Experimental Biology
Vol. 40, July 2002, pp. 780-784
Short-term androgen deprivation does not alter CaR and VDR mRNA expression
in duodenal mucosa in male rats
S Tiwari, S K Gupta, M Mehrotra, G Agarwal , P K Awasthi & M M Godbole
Received 4 December 2001; revised 26 February 2002
Androgen deprivation is associated with decline in intestinal calcium absorption. The effect of androgen on CaR and VDR intestinal
mucosa has not yet been studied. Calcium homeostasis, areal bone mineral density (aBMD, dual energy X-ray absorptiometry) and
expression of CaR and VDR mRNA in duodenal mucosa of orchidectomized (ORX) and sham operated (Sham) adult Sprague Dawley rats
at 4 week have been studied. There was no significant difference in serum calcium, alkaline phosphatase, calcidiol and calcitriol levels
between both the groups. Serum testosterone (T) (ng/dl) and inorganic phosphorous (iP) (mg/dl) levels were significantly lower in ORX
rats. As compared to sham rats, ORX rats had significant decline in in-vitro aBMD at proximal, middle and distal tibia, proximal, mid
and distal femur and femoral neck (P < 0.05). Northern blot analysis revealed no significant alteration in the CaR and VDR mRNA
expression in duodenal mucosa in ORX rats. CaR and VDR mRNA expression in duodenal mucosa is therefore, not affected by
physiological concentrations of testosterone in rats.
Indian Journal of Experimental Biology
Vol. 40, July 2002, pp. 785-790
Effect of phosphocreatine on H+ extrusion, pHi and dimorphism
in Candida albicans
Nikhat Manzoor, M Amin & Luqman A Khan*
Received 13 July 2001; revised 26 December 2001
Candida albicans is an opportunistic pathogen. Its proliferation in human hosts is believed to be controlled by
immunologic mechanisms. The plasma membrane of the fungus posseses an H+-ATPase (PM-ATPase) which actively
extrudes protons to generate an electrochemical gradient which is used in co-transport of nutrients. This ATPase is
associated with the growth, dimorphism and pathogenicity of the fungus. The physiological concentration of
phosphocreatine (PCr) is 20-35 mM in skeletal muscles. H+-extrusion in Candida cells was strongly inhibited by PCr; 44%
at 20mM and 69% at 40mM. H+-extrusion was stimulated 6.2-fold in the presence of 10mM glucose. This glucose
stimulated extrusion was inhibited significantly by PCr; 36% at 20mM and 53% at 40mM. The intracellular pH pattern of
cells destined to differentiate was greatly altered in the presence of PCr. Evagination time for control cells was between 90120 min. PCr, delayed dimorphism, reduced the population of cells differentiating to hyphae and also reduced the length of
hyphae after each time interval. Only 60% differentiation was observed with 10mM PCr and 40% for higher PCr
concentration even after 210 min. Direct interaction of PM-ATPase and PCr has been demonstrated by difference spectrum
measurement employing stopped flow spectrophotometer. It can be concluded that PCr may be playing a significant role in
checking growth and pathogenesis of C. albicans.
Indian Journal of Experimental Biology
Vol. 40, July 2002, pp. 791-795
Control of hyperglycemia and retardation of cataract by mulberry
(Morus indica L.) leaves in streptozotocin diabetic rats
B Andallu
and
N Ch Varadacharyulu
Received 18 June 2001; revised 21 December 2001
Dried leaf powder of mulberry (M. indica L.) when given along with the diet at 25% level to streptozotocin induced
diabetic male Wistar albino rats for 8 weeks, controlled hyperglycemia, glycosuria, albuminuria and retarded onset of
retinopathy. Untreated diabetic rats showed hyperglycemia, glycosuria, albuminuria and developed lenticular opacity after 8
weeks of experimental period.
Indian Journal of Experimental Biology
Vol. 40, July 2002, pp. 796-801
Growth behaviour and bioproduction of indole acetic acid by a Rhizobium sp.
isolated from root nodules of a leguminous tree Dalbergia lanceolaria
A C Ghosh & P S Basu
Received 28 February 2001; revised 5 February 2002
The Rhizobium sp. isolated from healthy and mature root nodules of a leguminous tree, Dalbergia lanceolaria Linn.f.,
preferred mannitol and KNO3 for growth as carbon and nitrogen sources, respectively. The bacterium produced a high
amount (22.3 g/ml) of indole acetic acid (IAA) from L-tryptophan supplemented basal medium. Growth and IAA
production started simultaneously. IAA production was maximum at 20 hr when the bacteria reached the stationary phase of
growth. Cultural requirements were optimized for maximum growth and IAA production. The IAA production by the
Rhizobium sp. was increased by 270.8% over control when the medium was supplemented with mannitol (1%,w/v), SDS
(1g/ml), L-asparagine (0.02%,w/v) and biotin (1g/ml) in addition to L-tryptophan (2.5 mg/ml). The possible role of IAA
production in the symbiosis is discussed.
Indian Journal of Experimental Biology
Vol. 40, July 2002, pp. 802-806
Secretion of ligninperoxidase by Penicillium citrinum, Fusarium oxysporum
and Aspergillus terreus
Meera Kumari, R S S Yadav & K D S Yadav
Received 8 May 2001; revised 12 February 2002
Secretion of ligninperoxidase [E.C.1.11.1.7] by Penicillium citrinum, Fusarium oxysporum and Aspergillus terreus in
liquid culture growth medium has been demonstrated. Enzymatic characteristics like K m , pH and temperature optima using
veratryl alcohol as the organic substrate of ligninperoxidases from above sources have been determined. Km values using
veratryl alcohol as substrate for enzymes from P. citrinum, F. oxysporum and A. terreus were 69, 64 and 60µM respectively.
Km values using H2O2 as the variable substrate were 64, 72 and 80µM.The pH optima were 4.0, 2.3 and 2.0 respectively.
The values of temperature optima were 30°, 25° and 22°C for the enzymes from P. citrinum, F. oxysporum and A. terreus
respectively.
Indian Journal of Experimental Biology
Vol. 40, July 2002, pp. 807-811
Effect of low level exposure of lead and cadmium on hepatic estradiol metabolism
in female rats
Anil Pillai, Laxmipriya, Ami Rawal & Sarita Gupta
Received 9 April 2001; revised 8 February 2002
Toxic effect of metal cations on female reproduction and gonadal functions was studied. Adult synchronized female
rats were treated intraperitoneally with lead acetate and cadmium acetate separately and in combination (0.025, 0.05 and 0.1
mg/kg body wt) for 15 days. The metabolizing enzymes (17-hydroxy steroid oxidoreductase and UDP glucoronyl
transferase) activities decreased with increasing dose showing significant change compared to control. Also, significant
decrease in cytochrome P450 (CYP450) content was found after the treatment. Displacement of zinc bound to
metallothionein was more in cadmium treated rats compared to other groups. In all these parameters, treatment in
combination of lead and cadmium showed intermediate results indicating some kind of competition between the two metals.
But the histological studies showed that combined treatment caused more cytotoxic effect than cadmium and lead alone.
These results indicated that metal cations tested did have a direct inhibitory effect on metabolizing enzyme activities.
Indian Journal of Experimental Biology
Vol. 40, July 2002, pp. 812-827
Screening of Indian plants for biological activity*: Part XVI
A K Goel, D K Kulshreshtha, M P Dubey & S M Rajendran
Received 15 October 2001; revised 16 January 2002
Alcoholic extracts of 288 of plant materials from 199 plant species have been tested for various biological activities
including chemotherapeutic and pharmacological screening. Biological activities, ranging from moderate to good degree,
have been observed in 61 plants extracts. Follow up studies have been carried out in these extracts and some of them have
shown moderate degree of activities at this Institute. However, none of the extracts was found to be good enough for further
development. Results of the present studies, alongwith chemical investigations on different species of similar genera which
were screened earlier, are also discussed.
Notes
Indian Journal of Experimental Biology
Vol. 40, July 2002, pp. 828-830
Antimicrobial potentiality of a phenothiazine group of antipsychotic
drug-prochlorperazine
Rupa Mazumder, Sharmila Ray Chaudhuri & Avijit Mazumder
Received 26 April 2001; revised 11 March 2002
The antipsychotic drug, prochlorperazine (Pcp), was tested for its antimicrobial efficacy against 103 strains belonging
to both gram positive and gram negative bacteria. The drug was found to possess maximum activity against Staphylococcus
aureus, Vibrio cholerae and Shigella spp. Pcp was moderately active against E.coli but most of the strains belonging to
Bacillus spp, Klebsiella spp, Salmonella spp and Lactobacillus spp were found to be resistant to this drug. The drug was
tested for its mode of antibacterial activity against Shigella dysenteriae 1 and it was found to be bacteriostatic in action. In in
vivo studies, Pcp offered significant protection to Swiss albino mice at concentrations of 0.75 g/g (P<0.01) and 1.5 g/g
(P<0.001) body weight when challenged with 50 median lethal dose of Salmonella typhimurium NCTC 74. Thus the result
depicts that prochlorperazine may emerge as a strong antimicrobial drug to replace the conventional antibiotics and to
overcome the problem of drug resistance.
Indian Journal of Experimental Biology
Vol. 40, July 2002, pp. 831-834
Synergistic effect of ayurvedic pearl preparation on enhancing effectiveness
of antibiotics
Manisha Kulkarni, J Y Deopujari & H J Purohit
Received 19 November 2001; revised 12 February 2002
Studies were carried out with ayurvedic preparations derived from pearl, which include preparations bhasma and pishti.
The synergistic effect to reduce the dose of antibiotic was tested against E .coli the test bacterium with ampicillin antibiotic
by bore well and disks diffusion methods. It was observed that pearl preparations do not show any antibacterial activity but
when used at 200 g/ml concentration with antibiotics, then even at sub-lethal dose, the antibiotic has effectively shown the
results with reduced contact time. The protocol was also tested with the other bacteria like, Pseudomonas aeruginosa, Vibrio
cholarae, Salmonella typhi, and Staphylococcus aureus and has shown similar results. The pearl bhasma synergistic effect
was also tested with other antibiotics such as erythromycin, kanamycin, and ampicillin.
Indian Journal of Experimental Biology
Vol. 40, July 2002, pp. 835-838
SEM study on cytotoxic effect of monocrotophos (MCP) on lungs of rat
Sangeeta, S M Handa & P K Mittal
Received 23 November 2001; revised 19 February 2002
Monocrotophos (MCP) on oral administration (0.28 mg/100 g of body wt. i.e. 1/5 th of LD50) to female rats for 15 and
30 days damaged alveolar walls lined by type II cells (great alveolar cells); clara cells (non-ciliated cells) lining bronchiolar
epithelium; and emphysematous lesions due to loss of inter-alveolar walls. This led to increase in surface tension in lung due
to decrease in secretion of surfactant as a result of necrosis of great alveolar cells and clara cells resulting in hypoxia. This
effect was time dependent. In R group (15 days without pesticide after 30 days daily oral treatment), the toxic effects
mentioned above still persisted which revealed non-repair of necrosis caused by MCP.
Indian Journal of Experimental Biology
Vol. 40, July 2002, pp. 839-841
Use of RAPD in assessing the genetic variability in Spodoptera litura
S Janarthanan, S Seshadri, K Kathiravan & S Ignacimuthu
Received 20 July 2001; revised 19 February 2002
A polymerase chain reaction based assay to distinguish six different ecotypes of the Spodoptera litura, a sporadic insect
pest and a predominant defoliator of various crops was carried out. A total of 40 random primers were screened to reveal the
existence of polymorphism between the populations. Among them eight showed scorable banding patterns and three primers
(OPA-01, OPA-05, OPM-01) exhibited distinguishable banding patterns. However, Chengalpattu and Chennai populations
revealed their closed relatedness and Coimbatore population stood distantly from others.
Indian Journal of Experimental Biology
Vol. 40, July 2002, pp. 842-845
Entomopoxvirus of cotton bollworm, Helicoverpa armigera (Hbn.)
K Narayanan
Received 10 September 2001; revised 16 November 2001
Occurrence of an Entomopoxvirus (EPV) from a lepidopteran insect viz;. cotton bollworm, H. armigera (HaEPV)
along with gross pathological symptoms is reported for the first time in India. Histopathological study revealed that the fat
body being the most favoured site of infection followed by haemocytes and gut epithelium. HaEPV was found to be not
cross infective to six of the agricultural lepidopteran insect pests except for the potato black cutworm, Agrotis segetum
registering 100% mortality showing typical symptom. Further, safety of HaEPV was shown against beneficial insect like
mulberry silkworm, Bombyx mori and an useful insect general predator, Chrysoperla carnea.
Indian Journal of Experimental Biology
Vol. 40, July 2002, pp. 846-849
Mass production of polyhedral occlusion bodies of NPV of Helicoverpa armigera
in relation to dose, age and larval weight
K Narayanan
and
S Jayaraj
Received 8 August 2001; revised 16 November 2001
A significant difference was noticed in the yield of polyhedral occlusion bodies (POBs) in various larval instars of H.
armigera when three different doses of the nuclear polyhedrosis virus (NPV) were administered. The yield of POBs from a
single larva ranged from 0.35  106 to 25033.33  106 with a mean of 18422.33  106 for fourth instar inoculated. Positive
correlation existed between larval weight and number of POBs recovered. The regression analysis indicated POBs recovered
responded with predictable manner to the weight of different larval instars and the various concentration of virus
administered. The medium lethal time increased in the instars of the larva advanced with a minimum of 3.5 and maximum of
8 days in the first and fifth instars respectively.
Indian Journal of Experimental Biology
Vol. 40, July 2002, pp. 850-853
In vitro regeneration of Acacia catechu Willd. from callus and mature
nodal explants  An improved method
Manisha Thakur, D R Sharma, Kamlesh Kanwar & Anil Kant
Received 6 September 2001; revised 5 February 2002
Callus was derived from cultured cotyledons on MS medium supplemented with 2,4-D (0.25 mg/l) and NAA (0.25
mg/l). Plantlets were regenerated from the callus and nodal explants on MS medium containing BAP (2.0 mg/l) and Kn (2.0
mg/l), and further multiplied on the same medium. Addition of adenine sulphate (25.0 mg/l), ascorbic acid (20.0 mg/l) and
glutamine (150.0 mg/l) in the medium resulted in enhanced axillary branching. Multiple shoots formed after 6 weeks were
separated and subcultured in the fresh medium of same composition. For rhizogenesis, microshoots of 2.0-2.5 cm length
were dipped in sterilized IAA solution (10 mg/l) for 24 hr followed by transfer to half strength MS medium containing
activated charcoal (0.02%) resulting in rooting (75%) within 8 weeks. The rooted plants were transferred to pots containing
sterilized soil and sand mixture for hardening and 71% survival was recorded. Fifty true to type plantlets of A. catechu could
be obtained within seven months of culture establishment.
Indian Journal of Experimental Biology
Vol. 40, July 2002, pp. 854-857
Cyanobacterial N2 fixation in presence of nitrogen fertilizers
A E Mekonnen, Radha Prasanna & B D Kaushik
Received 8 August 2001; revised 21 March 2002
Anabaena oryzae ARM 570 was examined for its growth (chlorophyll and protein), heterocyst frequency, nitrogenase
(acetylene reduction) activity, ammonia excretion, and glutamine synthetase and nitrate reductase in response to two levels
of urea-N vis-à-vis N2-N. Growth of cyanobacterium increased with duration of incubation. Reduction in heterocyst
frequency (40%) was observed at 30 ppm of urea-N, whereas at 60 ppm of urea-N, filaments were completely devoid of
heterocysts and no nitrogenase activity was observed. Maximum excretion of ammonia occurred at 30 ppm of urea-N, which
was concomitant with minimum glutamine synthetase activity. These results suggested that A. oryzae could be effectively
utilized in cyanobacterial biofertilizer programme even in the presence of combined nitrogen, for improving N-budget in
rice cultivation.
Indian Journal of Experimental Biology
Vol. 40, July 2002, pp. 858-860
Effect of lead on Na+, K+-ATPase activity in Penaeus indicus postlarvae
C Satyavathi & Y Prabhakara Rao
Received 22 June 2000; revised 9 January 2002
In vivo effect of lead on Na+, K+-ATPase was studied in plasma membrane/mitochondrial fraction of P. indicus
postlarvae (PL), exposed to 30 days to a sublethal concentration (1.44 ppm) of lead. A significant (P < 0.05) decrease in the
enzyme activity was observed for exposed PL with respect to their controls at different intervals except 24hr. Further the
substrate (ATP) and ion (Na+ and K+)-dependent kinetics of Na+, K+-ATPase was studied with the plasma
membrane/mitochondrial fractions of control and 30 days exposed PL. The apparent K M and Vmax values were calculated to
determine the nature of inhibition. Both the control and exposed PL showed almost the same apparent K M values in the
presence of different substrate or ion concentrations indicating that lead interacts with the enzyme at a different binding site.