The Aedes aegypti mosquito
Find information on thousands of medical conditions and prescription drugs.

Chikungunya

Chikungunya is a rare form of viral fever caused by an alphavirus that is spread by mosquito bites from the Aedes aegypti mosquito. The name is derived from the Swahili word meaning "that which bends up" in reference to the stooped posture developed as a result of the arthritic symptoms of the disease. more...

Home
Diseases
A
B
C
Angioedema
C syndrome
Cacophobia
Café au lait spot
Calcinosis cutis
Calculi
Campylobacter
Canavan leukodystrophy
Cancer
Candidiasis
Canga's bead symptom
Canine distemper
Carcinoid syndrome
Carcinoma, squamous cell
Carcinophobia
Cardiac arrest
Cardiofaciocutaneous...
Cardiomyopathy
Cardiophobia
Cardiospasm
Carnitine transporter...
Carnitine-acylcarnitine...
Caroli disease
Carotenemia
Carpal tunnel syndrome
Carpenter syndrome
Cartilage-hair hypoplasia
Castleman's disease
Cat-scratch disease
CATCH 22 syndrome
Causalgia
Cayler syndrome
CCHS
CDG syndrome
CDG syndrome type 1A
Celiac sprue
Cenani Lenz syndactylism
Ceramidase deficiency
Cerebellar ataxia
Cerebellar hypoplasia
Cerebral amyloid angiopathy
Cerebral aneurysm
Cerebral cavernous...
Cerebral gigantism
Cerebral palsy
Cerebral thrombosis
Ceroid lipofuscinois,...
Cervical cancer
Chagas disease
Chalazion
Chancroid
Charcot disease
Charcot-Marie-Tooth disease
CHARGE Association
Chediak-Higashi syndrome
Chemodectoma
Cherubism
Chickenpox
Chikungunya
Childhood disintegrative...
Chionophobia
Chlamydia
Chlamydia trachomatis
Cholangiocarcinoma
Cholecystitis
Cholelithiasis
Cholera
Cholestasis
Cholesterol pneumonia
Chondrocalcinosis
Chondrodystrophy
Chondromalacia
Chondrosarcoma
Chorea (disease)
Chorea acanthocytosis
Choriocarcinoma
Chorioretinitis
Choroid plexus cyst
Christmas disease
Chromhidrosis
Chromophobia
Chromosome 15q, partial...
Chromosome 15q, trisomy
Chromosome 22,...
Chronic fatigue immune...
Chronic fatigue syndrome
Chronic granulomatous...
Chronic lymphocytic leukemia
Chronic myelogenous leukemia
Chronic obstructive...
Chronic renal failure
Churg-Strauss syndrome
Ciguatera fish poisoning
Cinchonism
Citrullinemia
Cleft lip
Cleft palate
Climacophobia
Clinophobia
Cloacal exstrophy
Clubfoot
Cluster headache
Coccidioidomycosis
Cockayne's syndrome
Coffin-Lowry syndrome
Colitis
Color blindness
Colorado tick fever
Combined hyperlipidemia,...
Common cold
Common variable...
Compartment syndrome
Conductive hearing loss
Condyloma
Condyloma acuminatum
Cone dystrophy
Congenital adrenal...
Congenital afibrinogenemia
Congenital diaphragmatic...
Congenital erythropoietic...
Congenital facial diplegia
Congenital hypothyroidism
Congenital ichthyosis
Congenital syphilis
Congenital toxoplasmosis
Congestive heart disease
Conjunctivitis
Conn's syndrome
Constitutional growth delay
Conversion disorder
Coprophobia
Coproporhyria
Cor pulmonale
Cor triatriatum
Cornelia de Lange syndrome
Coronary heart disease
Cortical dysplasia
Corticobasal degeneration
Costello syndrome
Costochondritis
Cowpox
Craniodiaphyseal dysplasia
Craniofacial dysostosis
Craniostenosis
Craniosynostosis
CREST syndrome
Cretinism
Creutzfeldt-Jakob disease
Cri du chat
Cri du chat
Crohn's disease
Croup
Crouzon syndrome
Crouzonodermoskeletal...
Crow-Fukase syndrome
Cryoglobulinemia
Cryophobia
Cryptococcosis
Crystallophobia
Cushing's syndrome
Cutaneous larva migrans
Cutis verticis gyrata
Cyclic neutropenia
Cyclic vomiting syndrome
Cystic fibrosis
Cystinosis
Cystinuria
Cytomegalovirus
Dilated cardiomyopathy
Hypertrophic cardiomyopathy
Restrictive cardiomyopathy
D
E
F
G
H
I
J
K
L
M
N
O
P
Q
R
S
T
U
V
W
X
Y
Z
Medicines

This disease was first described in Tanzania, Africa in 1952. An outbreak of Chikungunya was discovered in Port Klang in Malaysia in 1999 affecting 27 people .

A serological test for Chikungunya is available from the University of Malaya in Kuala Lumpur, Malaysia.

It is closely related to O'nyong'nyong virus. (PMID 15891138)

Symptoms

The symptoms of this disease include fever which can reach 39 °C, a rash typically described as petechial or maculopapular usually involving the limbs and trunk, and arthralgia or arthritis affecting multiple joints which can be debilitating in severity. There can also be headache, conjunctival injection and slight photophobia.

Treatment

There is no specific treatment for Chikungunya. The illness is usually self-limiting and will resolve with time. Symptomatic treatment is recommended after excluding other more dangerous diseases.

Read more at Wikipedia.org


[List your site here Free!]


Insect cell culture in research: Indian scenario
From Indian Journal of Medical Research, 6/1/05 by Sudeep, A B

Insect cell cultures are widely used in viral diagnosis and biotechnology, for the production of recombinant proteins, viral pesticides and vaccines as well as in basic research in genetics, molecular biology, biochemistry, endocrinology and virology. Following KRP Singh's pioneering research in 1967, a large number of cell lines from diptera, hemiptera, and lepidopteran insects were established and characterized in India. With the availability of the modern tools in molecular biology and the advancements made in biotechnology, the indigenous cell lines may prove useful in creating a future without biohazardous chemical pesticides as well as producing life saving pharmaceuticals and vaccines for many diseases. This review summarizes information gathered regarding the insect cell lines established so far in India. It also covers the familiarization of the well characterized continuous cell lines and their potential applications. Special attention is given to virus susceptibility of the cell lines, the yield of virus with a comparative analysis with other conventional systems. The potential applications of dipteran and lepidopteran cell lines in agriculture and biotechnology are also briefly discussed for prospective studies.

Key words

Baculoviruses - dipteran and lepidopteran cell lines - insect cell culture - recombinant proteins

The successful demonstration of growth of fibrillae from embryonic frog neuroblasts in clotted lymph by Harrison1 and subsequent workers has opened up a new scientific field of animal tissue culture. The importance achieved in a short span in vertebrate cell culture research has warranted an urgent need to have a parallel system from arthropods especially from insects. Arthropods have become the centre of attraction of scientists due to their importance in medicine and agriculture. They played an important role as intermediate hosts or vectors of pathogens causing diseases to man and pet animals. The growing interest in arbovirus infections of man and domestic animals has stimulated interest in developing cell cultures from arthropods of medical importance especially from mosquitoes and ticks in order to study the vector virus relationship at cellular level. Similarly, in the field of agriculture, the damage caused by caterpillars and the failure of insecticides in their control stimulated urgency in developing cell cultures from lepidopteran insects for large-scale production of insect viruses. The first breakthrough in the development of insect cell culture was the report of maturation of testicular follicle cells to spermatocytes in Cercopia moth in an artificial medium2. This result gave encouragement to many workers to initiate work on the development of insect cell culture3-5; however, most of these earlier attempts were disappointing6.

The basic reason for the failure was the lack of a suitable cell culture medium for culturing insect cells. Efforts were made in the late 1940s and 1950s for a medium whose composition resembled that of insect hemolymph. The first such medium was formulated by Wyatt which proved useful for cultivating silk moth ovarian tissues7. Subsequently many formulations, which gave promising results, were reported8-11. However, the establishment of a viable self-sustainable insect cell line was not possible. Nonavailability of a nutritionally optimum growth medium to stimulate multiplication of cells in vitro, difficulty in obtaining adequate quantity of sterile tissues as well as the lack of adequate knowledge of tissue culture techniques etc., could have been the possible reasons for this. However, a few promising results appeared in the late 1950s12-13 though these studies did not culminate in the establishment of continuous cell lines. Vago and Chastang12 cultivated the larval ovary cells of Bombyx mori up to the 12th passage level while Gaw and colleagues13 subcultured the epithelial cells of B. mori gonads up to the 22nd passage level. These studies indicated the probability of growing insect cells in vitro for the first time that have stimulated further research.

The first true cell line from an arthropod was established from the pupal tissues of the moth Antherea eucalypti by TDC Grace of Australia in 1962(14). He modified Wyatt's medium7 with the addition of 10 vitamins and formulated a new medium known as Grace's insect tissue culture medium, which supported the growth of the cell line. This medium became popular due to its usefulness in the establishment and maintenance of many insect cell lines. Today, this formulation is available commercially and several workers have modified this medium to suit their demands15,16. The study of insect cells made rapid progress in the following years and more than 500 cell lines from different insect species have been described till today17. The system has found wide application in the studies on morphogenesis, virology, pathology, biochemistry, genetics and other fields of biology and medicine.

In recent years, there is renewed interest in developing new lepidopteran cell lines due to their potential application in biotechnology. The advances made in the genetic engineering technology in the early eighties have helped in the production of new recombinant proteins and genes useful in medicine and agriculture18,19. Lepidopteran cell lines were primarily being established to propagate insect viruses as a biopesticide for the control of insect pests. Lately, the baculovirus expression vector system combined with insect cell cultures has become more attractive for the expression of many heterologous proteins than other systems viz., bacterial, yeast, vertebrate viruses, etc., due to its unique characteristics. This technology is also being used in the construction of recombinant baculoviruses to use as biopesticides that offered comparatively faster killing of insect pests than by wild type baculoviruses20-23.

Mosquito cell lines

Mosquitoes transmit a number of diseases to human beings and domestic animals. The importance of mosquitoes as vectors of many protozoan, viral and filarial pathogens is well documented. Efforts to control the mosquitoes have not been very successful due to their ability to adapt to different environmental conditions as well as to develop resistance to synthetic insecticides24. Susceptibility of mosquitoes to these pathogens is a very complex phenomenon, which is governed by several genes and environmental conditions. The availability of established mosquito cell lines came handy for understanding the interaction of pathogens at the cellular level. The available information indicated that attempts to develop mosquito cell lines have started in the early 1930s3.

Grace in 1966 reported the establishment of the first mosquito cell line in the world from Aedes aegypti mosquitoes25. Subsequent studies using isoenzyme analysis indicated this cell line as a contaminant of Antherea eucalypti cell line26. Singh27 established two cell lines at the National Institute of Virology (NIV), Pune from the larval tissues of Ae. aegypti and Ae. albopictus in Mitsuhashi Maramorosch (MM) medium, which was originally designed for culturing leaf hopper cells by Mitsuhashi and Maramorosch28. Singh's cell lines are therefore considered as the first, true mosquito cell lines established in the world. He solved the problems associated with obtaining sterile tissues for initiation of cell cultures using a simple technique for surface sterilization of mosquito eggs. In addition, he used commercially available foetal bovine serum (FBS) as a supplement in place of insect haemolymph, which was considered essential for establishment and maintenance of mosquito cell cultures. Subsequently, a number of mosquito cell lines from different species were established in India and abroad. Table I depicts the number of mosquito cell lines established at NIV, Pune, India.

Attempts were made in the early 1980s to develop cell lines from Culex mosquitoes due to their importance in the transmission of Japanese encephalitis (JE) virus in India. The virus has established itself in several parts of India and became a major cause of concern35. The first Culex cell line in the country was established from the embryonic tissue of C. bitaeniorhynchus mosquitoes32. This species is considered as one of the important vectors of JE in India35,36 and Banerjee et al31 have shown the transmission of JE virus by this mosquito in the laboratory. Cell lines from other two species of Culicine mosquitoes viz., C. infula, C. ambiguous, which are members of C. bitaeniorhynchus complex were also established subsequently38. Among these cell lines, the C. bitaeniorhynchus cell line was found highly susceptible to JE and West Nile (WN) viruses. The cell line also supported the multiplication of other arboviruses of public health importance in India viz., Chikungunya (CHIK), dengue (DEN) and Sindbis viruses39. This cell line was also used in the epitope analysis of different strains of JE virus using monoclonal antibodies40.

A new cell line from the forest mosquito Ae. krombeini was established in the early 1990s, which proved highly sensitive to several arboviruses33. The new cell line supported the replication of 13 mosquito borne arboviruses41'42. It was extremely sensitive to JE and DEN viruses and very low titre of these viruses could be detected on the 1st post infection day (PID) using indirect immunofluorescent technique (IFA) while C. bitaeniorhynchus and C6/36 cell lines could not detect these viruses when infected with same dilution43. Unfortunately, due to the presence of certain cytoplasmic inclusions in the cells44, the cell line was not used for further research especially for isolation of virus from field collected mosquitoes.

Recently, a new cell line from C. tritaeniorhynchus mosquitoes was established from the embryonic tissue34. This mosquito, an important member of the C. vishnui complex, is the main vector of JE virus in India35,45. The cell line supported the multiplication of JE and WN viruses but did not support the multiplication of any of the dengue serotypes34.

Applications of mosquito cell lines in virus research

Most of these cell lines are interesting from an academic point of view and much of the early research work was concerned with characterizing the basic physiology of cells in culture. The development of continuous cell lines has proved useful in diverse areas of research especially in virology. The arbovirus-arthropod cell relationships have been the prime consideration due to the subsequent application of mosquito cells in various aspects of arboviral research. Singh's Ae. albopictus cell line, which is popularly known as ATC-15 cell line was the most widely studied cell line due to its susceptibility to nearly 50 viruses transmitted by mosquitoes, ticks etc46. Singh's other cell line derived from Ae. aegypti (ATC-10) mosquitoes was less susceptible to arboviruses compared to ATC-15. Singh also observed that with the exception of Colarado tick fever and Ganjam viruses, those arboviruses that are naturally transmitted by mosquitoes or infect mosquitoes experimentally, multiplied in one or both of his cell lines. Extending this argument, Buckley also reported that partially deoxycholate resistant viruses can multiply in one of these cell lines where as lipid solvent sensitive arboviruses isolated from ticks and sand flies cannot multiply in either cell lines47.

(i) In primary isolation of arboviruses: The availability of continuous cell lines has been very useful in the study of arboviruses. The absence of cytopathic effect (CPE) in majority of mosquito cell lines infected with arboviruses has limited the use of this cell system for primary isolation work. However, several successful attempts have been made using those cell lines, which showed CPE for virus isolation from original field materials. Using ATC-15 cell line, Singh and Paul48 successfully demonstrated the isolation of four serotypes of DEN viruses from human sera. In their later studies, Paul and Singh49 compared the sensitivity of ATC-15 cell line, VERO cells and intracerebral inoculation of mice to some arboviruses and showed the superiority of the mosquito cell culture system. The results indicated that Ae. albopictus cell line was equally or slightly more sensitive to infection with CHIK, WN and JE viruses comparing the other two systems. However, with DEN-2 virus, the mosquito cell culture system was 100 and 40 times more sensitive than VERO and infant mice system, respectively. Supporting results were also published by Chappel and coworkers in the isolation of dengue viruses over infant mice and LLC-MK2 cells50.

IFA has become an important tool in the detection of virus antigen with its simplicity and specificity51. This technique is now routinely being used in the detection of virus antigen in infected cell lines and mosquitoes42,52. This new technique has helped to rediscover the potential of mosquito cell lines in the study of arboviruses, and several virus isolations were made in mosquito cell lines from field collected mosquitoes at NIV, Pune53-56. The C. bitaeniorhynchus cell line for JE virus and Ae. albopictus cell line or C6/36 cells57 for DEN viruses are mainly being used in the isolation of these viruses from mosquitoes collected during epidemic periods.

(ii) In serology: The use of mosquito cell lines in the serological typing of arboviruses has been restricted due to the lack of CPE in these cell lines. However, several successful attempts have been made to use those cell lines, which showed CPE58-63. A few studies were also reported from India. Paul and coworkers blocked the production of CPE in JE infected cultures with JE virus antiserum in Ae. albopictus cell line64, In another study, Pavri and Ghosh65 successfully used tissue culture fluid from DEN infected Ae. albopictus cells directly in complement fixation (CF) test. Subsequently, Singh and Paul followed this method for serotyping of dengue viruses48. Ghosh and associates66 observed CF antigens of Chandipura (CHP) and JE viruses in infected Ae. albopictus and Ae. aegypti cells. In JE virus infected cells, the CF antigen was found in both intra- and extra-cellular fluid whereas in CHP infected cells CF antigen was retained intracellularly. Ghosh and Bhat67 also reported haemagglutinin activity in normal culture fluid of Ae. albopictus cells.

(iii) In rickettsial studies: The replication of Coxiella burnetti was detected on the 7th PID in C. bitaeniorhynchus cell line using IFA68. The rupturing of cell walls and subsequent release of the organism into the tissue culture fluid (TCF) was also reported by the authors. This TCF was effectively used to infect embryonated chicken eggs. The IFA technique has proved quicker in detecting the presence of rickettsiae as in other conventional methods like staining with Gimanez staining, etc., the detection was possible only on the 11th PID.

(iv) In biochemical and molecular studies: In India, though many mosquito cell lines are available, these were used mainly in the isolation of arboviruses except for a few other studies69. Mosquitoes gained considerable importance in biochemical and genetic research due to their increasing resistance to insecticides. Following the pioneering work of Stollar and coworkers70, mosquito cell lines are routinely being" used to carry out genetic and biochemical aspects71-73. This system has been employed effectively in various fields of molecular biology and genetics in recent years especially to explore the suitability of established mosquito cell lines for investigating cellular and humoral immunity73. The cell cultures are also being used in the exploration of immune responses in mosquitoes, receptor studies, host parasite interaction, etc., in addition to their various other applications in molecular biology. Better results can be achieved by generating highly susceptible cell lines or cell clones, which show little or very low cellular responses against a particular pathogen.

Lepidopteran cell lines

In the last two decades, a lot of work has been done in India to develop cell lines from lepidopteran insects due to their potential application in the fields of agriculture and medicine. The cell lines are used as a tool to grow many entomopathogenic viruses, which have the potential as biopesticides. These viruses are highly virulent to susceptible insect hosts and are ecofriendly. Members of the family baculoviridae, in particular, have been projected as bio-pesticides of the future and are being used in many countries for pest control74. Baculoviruses have been isolated from the members of Phylum Arthropoda (insects and crustaceans) and no isolations have been reported from vertebrates so far75. Baculoviruses are extremely virulent to their susceptible hosts and are safe for humans. In nature, baculoviruses play an important role in bringing down the pest population to economic thresholds as and when their population increases20. In the field of medicine, the cell lines play an important role in the abundant expression of many life saving molecules, which are otherwise difficult to synthesize. The application of baculovirus expression vector system (BEVS) for the expression of foreign proteins in insect cells has stimulated tremendous interest in the recent years. The following unique characteristics distinguish BEVS from other systems and a wide variety of genes and medically important proteins from viruses, fungi, bacteria, plants and animals have been expressed in insect cells using BEVS76,77. These biomolecules are used effectively in pharmaceuticals and in the development of vaccines and diagnostics.

(i) Abundant expression of soluble recombinant . proteins, which are antigenically, immunologically and functionally similar to their native counter parts. (ii) Ability to accommodate large DNA inserts, (iii) Two or more proteins can be expressed simultaneously by using dual expression vectors or by coinfection with two or more different recombinant viruses, (iv) Baculovirus infected insect cells perform many of the post-translational modifications such as phosphorylation, glycosylation, correct signal peptide cleavage etc. (v) Very late transcription from the polyhedrin promoter allows expression of cytotoxic recombinant gene products, (vi) Baculoviruses and the cell system are easy to handle and safe.

In India, though NIV scientists were the pioneers in developing mosquito cell lines, not much attention was given to develop lepidopteran cell lines until recently. The establishment of the first lepidopteran cell line was reported by Pant et al in 1977(78). They established a cell line from the embryonic tissue of potato tuber moth (Gnorimoschema/Phthoramea operculella) and characterized the cell line in detail. The cell line was initiated and maintained in MM medium supplemented with 10 per cent FBS which was later replaced with 0.8 per cent bovine serum albumin supplemented with yeastolate and lactalbumin hydrolysate. This is one of the very few insect cell lines maintained in growth medium without supplementing FBS. The cells could also be grown as suspension culture in conical flasks on a rotary shaker giving higher yield of cells. However, the authors did not study the susceptibility of the newly established cell line to any of the baculoviruses.

It took nearly two decades to establish another lepidopteran cell line in India. In the last decade, though many scientists at different laboratories actively worked on development of new lepidopteran cell lines, only NIV, Pune succeeded in the establishment of cell lines and reported eight new cell lines during 1997-2002 (Table II). In India, lepidopteran cell lines i.e., Sf-9, Sf-21, BMN, etc., are being used for various studies and all these cell lines are procured from abroad. The new, indigenously developed lepidopteran cell lines were characterized in detail using the routine parameters including their susceptibility to several baculoviruses. Some of the cell lines have already been proved useful in propagating baculoviruses due to their high susceptibility and virus yield while some expressed recombinant proteins with better efficiency than Sf-9 cells (NIV unpublished data). Further research in these directions may produce the full potential of these cell lines in expressing foreign proteins and genes.

(i) Establishment oflepidopteran cell lines: Cell lines were established from different organs/tissues i.e., embryonic, ovarian, fat body, testis, imaginai discs, dorsal vessel, etc., at different stages of development from lepidopteran insects. This is possible due to the comparatively larger size of these insects. The maximum number of cell lines has been established from the embryonic tissues followed by ovarian tissue17. In India, majority of lepidopteran cell lines established were from the ovarian tissue followed by embryonic tissue (Table II). The initiation of cell cultures from different tissues of lepidopteran insects is shown diagrammatically in the Fig.

(ii) Characterization of cell lines: The newly established cell lines were characterized using routine parameters i.e., morphology, karyology, growth curve, species specificity, etc., as done for all other established cell lines in the world. Morphological studies carried out for all the newly established lepidopteran cell lines indicated that epithelial-like cells dominated in most of the cell lines followed by fibroblast-like cells and a few giant or vacuolated cells. Karyological analysis indicated a typical lepidopteran pattern showing a large number of dot-like, darkly stained chromosomes in all the cell lines and their number ranged from 45 to over 300. For morphological and karyological studies, the method described by Schneider85 was used. Growth of the cell lines differed individually and ranged from 4- to 10-fold weekly. Species specificity of all the cell lines was determined using isoenzyme profile analysis and heteroduplex analysis. Isoenzyme profile analysis is an important parameter used in the characterization of a new cell line to rule out the chances of cross contamination from the existing cell lines using the method described by Tabachnik and Knudson86. Four enzymes i.e., lactate dehydrogenase, malate dehydrogenase, Glucose-6-phosphate dehydrogenase and isocitric dehydrogenase, were used in the present study. The enzyme profiles indicated the cell lines as distinct, new cell lines, and not a contaminant of any of the cell lines maintained in the laboratory. The heteroduplex analysis is a new molecular technique used to authenticate cell lines and used effectively to differentiate cell lines recently87,88. The 12S or 16S rRNA gene of the mitochondrial DNA was PCR amplified and run on a poly acrylamide gel along with the corresponding amplified fraction of DNA from parent insect. The appearance of a homoduplex indicated 100 per cent homology while heteroduplex indicated cross contamination. The analysis of all the cell lines gave homoduplex with their parent DNA indicating the cell lines as new, unique cell lines (Shouche et al, personal communication).

(iii) Susceptibility of the established cell lines to certain baculoviruses: Spodoptera litura cell lines-S. litura is an important agricultural pest in India causing serious damage to tobacco and vegetables. Three new continuous cell lines from larval ovaries (NIV-SU-992), pupal ovaries (NIV-SU-893) and larval haemocyte (NIV-SU-1095) were established and characterized in detail79-81. Susceptibility of these cell lines to certain baculoviruses was also studied and found susceptible to the homologous virus Spodoptera litura multiple nucleopolyhedrovirus (SpItMNPV) as well as Autographa californica multiple nucleopolyhedrovirus (AcMNPV). On infection with SpItMNPV, the three cell lines yielded 5.6, 5.3 and 3.0 x 10^sup 6^ occlusion bodies (OBs)/ml, respectively on the 7th PID89. With AcMNPV, the cell lines yielded 5.5, 3.0 and 1.96 x 10^sup 6^ OBs/ml, respectively. However, none of these cell lines supported the growth of Helicoverpa armigera single nucleopolyhedrovirus (HaSNPV), Bombyx mon multinucleopolyhedrovirus (BmMNPV) and potato tuber moth granulosis (PTM-GV). The susceptibility status of these cell lines to different baculoviruses is given in Table III.

Helicoverpa armigera cell lines'. H. armigera is a polyphagous insect causing damage to over a hundred species of plants especially cash crops i.e., cotton, vegetables and ornamental plants causing damage worth billions of dollars globally90. In India, this is a major pest of cotton, vegetables and oilseeds. Being a borer, the larvae remain inside the stem or fruits for most of the time; hence, the application of chemical insecticides (contact poison) does not make much impact in the control of the larvae. Therefore, efforts are being made to control this insect using baculoviruses i.e., HaSNPV.

Presently, the production of HaSNPV is made in the larval system, which is labour intensive and expensive. Therefore, efforts were made to establish cell lines from this insect for production of HaSNPV in large scale. Two cell lines from the H. armigera were developed i.e., from the larval haemocyte and embryonic tissue and designated as NIV-HA-1195 and NIV-HA-197, respectively. Both the cell lines were tested for their susceptibility to different baculoviruses (Table III). The NIV-HA-197 cell line was highly susceptible to HaSNPV, yielding a very high titre (2.88 x 10^sup 7^ NPV/ml) on the 10th PID83. The cell line was also susceptible to AcMNPV and SpItMNPV yielding high titres91. The HaSNPV grown in the cell culture was bioassayed in different instar larvae of H. armigera. The results indicated that the OBs produced in vitro were highly virulent to 2nd and 3rd instar H, armigera larvae causing cessation of feeding on the 2nd day and mortality in 6 days. This cell line is also growing in goat serum (GS) supplemented medium producing a comparable yield of OBs. These OBs were found more virulent than the former and 100 per cent mortality was observed within 5 days of infection to 2nd instar H. armigera larvae (Sudeep, unpublished data). Goat serum, being cheap and locally available will help in the large scale production of HaSNPV for use as a biopesticide in future.

Bombyx mori cell lines: B. mori cell lines are effectively being used in the expression of foreign proteins using BEVS77,84. In India, research work has been carried out with B. mori cell lines, (BMN, BM5, etc.) brought from abroad, for expression studies92,93. Efforts were therefore, made to establish cell lines from this important insect and two cell lines were established from the larval and pupal ovaries. The newly developed larval and pupal ovary cell lines were designated as NIV-BM-1296 and NIV-BM-197, respectively and characterized in detail84. Both the cell lines were susceptible to BmMNPV and AcMNPV. However, the cell lines did not support the multiplication of HaSNPV and SpItMNPV (Table III).

Potato tuber moth (Gnorimoschema/Phthorimaea operculella Zeller) cell line: PTM is an important pest of potatoes in India causing extensive damage to the crop. This pest attacks the crop in the field as well as in storage, and efforts to control this insect using conventional methods has not been satisfactory94. Release of parasitoids and predators has also not helped in the control of this insect and the damage is increasing steeply. Efforts are being made to control with granulovirus (GV) in many laboratories and agricultural universities but with limited success. Large scale production of GV in the insect (in vivo) is not feasible due to the small size of the insect. The growth of PTM-GV in a newly established PTM cell line was reported by Lery and coworkers in Egypt95. Efforts were made in our laboratory and an embryonic cell line (NIV-PTM-1095) was established from the moth and characterized including the susceptibility to PTM-GV (unpublished data). On infection with PTM-GV, a few virus-like particles were observed in the tissue culture fluid in electron microscopic studies89. The cell line did not support the replication of any of the other baculoviruses tested (Table III). However, subsequent studies with HaSNPV showed a few occlusion bodies in a few cells. In AcMNPV infected cultures also, the morphology of the cells was found significantly changed in comparison to the control cells, however, occlusion bodies were not observed (Sudeep, unpublished data).

(iv) Application of the newly established lepidopteran cell lines for research-Nucleopolyhedrovirus production: Two new cell lines, H. armigera embryonic cell line (NIV-HA-197) and S. litura larval ovary cell line (NIV-SL-992), produced very high yield of OBs on infection with respective homologous viruses. OBs grown in vitro were found highly virulent to 2nd and 3rd instar larvae causing mortality within 6 days post infection (PI). Due to their importance in the field of agriculture, efforts are being made to scale up the production of these baculoviruses. The HaSNPV yield in FBS supplemented medium was 28.8 x 10^sup 6^ OBs/ml on the 10th PID in the NIV-HA-197 cell line. Efforts were also made to clone this cell line to get high virus yielding cell populations and seven clones were developed. The clones were studied for their virus yielding properties and two clones were found better than the other clones in virus production. Both the cell clones i.e., cl-1 and cl-7, yielded >32 x 10^sup 6^ OBs/ ml on the 7th day PI in FBS supplemented medium. The clones were adapted to goat serum supplemented medium to reduce the production cost. One of the clones (cl-1) yielded 32.3 x 10^sup 6^ OBs per ml in GS supplemented medium on the 10th day PI (Sudeep, unpublished data). Bioassay of GS supplemented medium grown OBs caused 100 per cent mortality to 2nd instar larvae of H, armigera on the 5th PID. The GS adapted sub-line may be useful in the large scale production of HaSNPV for use as a biopesticide. GS is locally available and can be processed for cell culture use at a very negligible cost which will be a plus point for commercial competitiveness.

In expression of foreign proteins (recombinant proteins): In India, imported cell lines i.e., Spodoptera frugiperda cell line (Sf-9 and Sf-21), Trichoplusia ni and B. mon cell lines have been used to study the expression of different foreign proteins92,93,97. We have made an effort to express recombinant (AcMNPV) proteins in the newly developed cell lines. Preliminary. experiments with hepatitis C virus (HCV) core protein indicated significant level of expression in H. armigera and S. litura cell lines. Further studies with two other recombinant proteins viz., P24 and gp120 of HIV-1 indicated high level of expression of these proteins in NIV-HA-197 and NIV-SU-1095 cell lines. The expression of P24 was also not affected significantly by the absence of FBS in the cell culture. NIV-HA-197 and NIV-SU-1095 cell lines showed higher level of expression of gp120 protein of HIV-1 than that observed in Sf-9 cells. The effect of tunicamycin, which inhibits glycosylation of glycoproteins, was also studied in these cell lines. Tunicamycin affected the gp120 synthesis in all the five cell lines tested. However, the effect was significant in H. armigera (embryonic) cells and S. litura (haemocyte) cells.

(v) Miscellaneous studies: In addition to diptera and lepidoptera, attempts were made to develop cell lines from other Orders of Class Insecta. Bhat and Singh98 attempted to develop a cell line from the triatomine bug, Triatoma rubrofasciatus. Though three different cell types along with multicellular vesicles were observed in the culture, they degenerated after 30 days. Pant and coworkers99 successfully established three cell lines from the bed-bug, Cimex hemipterus (Hemiptera: Cimicidae) and characterized one cell line (NIVI-CH-440) in detail. The susceptibility of the cell line was studied to important arboviruses viz. JE, CHIK, DEN-2, etc., and the rickettsiae, C. burnettii. It was interesting to note that neither the viruses nor the rickettsiae multiplied in this cell line.

Prospective study

A number of well characterized, dipteran and lepidopteran cell lines are available with NIV but the potentials of these cell lines are not yet fully exploited. Mosquito cell lines have been used mainly in the isolation of arboviruses. These cell lines with the help of modern tools available in molecular biology may find application in solving many of the complex problems associated with mosquito borne diseases.

Lepidopteran cell cultures are increasingly being used in diverse areas of research i.e., in biotechnology, agriculture and medicine. Many lepidopteran cell lines are being used to propagate insect viruses in large scale and are being used as biopesticides. The successful application of HaSNPV has been reported from China, Australia and other parts of the world74,90. Similarly, Anticarcia gemmatalis nucleopolyhedrovirus is being used for the control of velvet bean caterpillar of Soya bean in Brazil for the last two decades74. The US environment protection agency has approved the use of six baculoviruses for application against pests of agriculture and forestry100. In this context, it is high time to think in this direction and recognize the potential baculoviruses for crop protection in India. As mentioned earlier, H. armigera and S. litura are important pests of agriculture in India and may be controlled by the respective baculoviruses. The production of these viruses are presently made in the in vivo system and the new cell lines can replace the production of virulent OBs in large scale at a competitive cost. Already goat serum adapted cell lines are available for baculovirus propagation, which produce comparative yield with better virulence (Sudeep, unpublished data). Integrating the existing facilities and proper application of potentials may make this true and India can have a future without hazardous chemical pesticides.

Preliminary results obtained in the expression studies indicate that H. armigera (NIV-HA-197) and S. litura (NIV-SU-1095) cell lines yield better expression of certain proteins than Sf-9 cells, which are used universally for expression studies101. The new cell lines therefore, have the potential for expressing heterologous proteins using AcMNPV as a vector. It is important to note that in the cell lines the yield of AcMNPV was much lower compared to the yield of homologous viruses. However, both the cell lines yielded better expression of foreign proteins in comparison to Sf-9 cells. It will be worth to construct recombinants in HaSNPV or SpItMNPV and try to express foreign proteins in these cell lines. Due to their high susceptibility to homologous baculoviruses, these may produce significantly high amount of recombinant proteins. The higher level of expression also contributes in reducing the production cost. Another cell line, which can be used for expression studies, is B. morl (larval ovary) cell line, which produced very high yield of BmMNPV. B. mori cell lines are extensively used in expression studies with comparable efficiency, and a number of foreign proteins were expressed using BmMNPV as vector77,84. The new cell lines developed in India have not been studied for expression of foreign proteins. However, due to their high susceptibility to BmMNPV, these cell lines may also express heterologous proteins with better efficiency.

In India, though we have developed a few lepidopteran cell lines there is an urgent need to develop more new indigenous cell lines, as these may find better application than the existing cell lines in their virus yielding properties. The unique features of the BEVS and the economic benefits in the production of proteins as reagents, diagnostics or therapeutics are important in the fields of medicine and agriculture suggest that this system has great potential for the future. The new cell lines along with the existing ones will definitely help in future research to develop biological pesticides as well as the large scale production of important life saving pharmaceutical products.

Acknowledgment

Authors thank Dr G. Geevarghese, and PVM Mahadev, for critically reading the text and also for valuable suggestions.

References

1. Harrison, RG. Observations on the living developing nerve fibre. Proc Soc Exp BM (NY) 1907; 4 : 140-3.

2. Goldschmidt, R. Some experiments on spermatogenesis in vitro. Proc Natl Acad Sd USA 1915; 1 : 220-2.

3. Trager W. Multiplication of virus of equine encephalomyelitis in surviving mosquito tissues. Am J Trop Med 1938; 18 : 387-93.

4. Garilov N, Cowez S. Essai de culture in vitro de tissue de mostiques et d' intestine de Lapina adult insects. Ann Parasitai Hung Comp 1941; 18: 180-6.

5. Carlson JG. Protoplasmic viscosity changes in different regions of the grasshopper neuroblasts during mitosis. Biol Bull Woods Hole 1946; 90 : 109-21.

6. Day MF, Grace TDC. Review of recent work on insect tissue culture. Ann Rev Entomol 1959; 4 : 17-38.

7. Wyatt SS. Culture in vitro of the tissue from the silkworm Bombyx mori. J Gen Physiol 1956; 29 : 841-52.

8. Grace TDC. Effects of various substances on growth of silkworm tissues in vitro. Aust J Exptl Biol 1958; 11 : 407-11.

9. Grace, TDC. The prolonged growth and survival of ovarian tissue of the promethea moth, Callosamia promethea, in vitro. J Gen Physiol 1958; 41 : 1027-34.

10. Jones BN, Cunningham I. Growth by cell division in insect tissue culture. Nature 1960; 187 : 1072-4.

11. Jones BN, Cunningham I. Growth by cell division in insect tissue culture. Exp Cell Res 1961; 23 : 386-401.

12. Vago C, Chastang S. Obtention de lignees cellulaires en culture de tissue d' invertebres. Experientia 1958; 14 : 110-1.

13. Gaw Z. Liv NT, Zia TU. Tissue culture method for cultivation of grasserie. Acta Virol 1959; 3 (Suppl): 55-60.

14. Grace, TDC. Establishment of four strains of cells from insect tissues grown in vitro. Nature 1962; 795 : 788-9.

15. Hink WF, Ellis BJ. Establishment and characterization of two new cell lines (CP-1268 and CP-169) from the codling moth, Carpocapsa pomonella (with a review of culture of cells and tissues from Lepidoptera). Curr Top Microbiol Immunol 1971; 55 : 19-28.

16. Mitsuhashi J. Development of highly nutritive culture media. In Vitro Cell Dev Biol (Animal) 2001; 37 : 330-7.

17. Lynn DE. Novel techniques to establish new insect cell lines. In Vitro Cell Dev Biol (Animal) 2001 ; 37 : 319-21.

18. Smith GE, Summers MD, Frazer MJ. Production of human beta interferon in insect cells infected with a baculovirus expression vector. MoI Cell Biol 1983; 3 : 2156-65.

19. Carbonell LF, Hodge MR, Tomalski MD, Miller MK. Synthesis of a gene coding for an insect-specific scorpion neurotoxin and attempts to express it using baculovirus vectors. Gene 1988; 73 : 409-18.

20. Wood HA. Development and testing of genetically improved baculovirus insecticides. In: Shuler ML, Wood HA, Granados RR, Hammer DA editors. Insect cell cultures: Production of improved bio-pesticides and proteins from recombinant DNA. New York: Wiley-Liss; 1995 p. 91-130.

21. Tracey MF, All JN, Glidia GM. Effect of ecdysteroid UDP-glycosyl transferase gene deletion on efficacy of a baculovirus against Heliothis virecens and Trichoplusia ni (Lepidoptera: Noctuidae). J Econ Entomol 1997; 90 : 1207-14.

22. Gershberg E, Stockholm D, Froy O, Rashi S, Gurevitz M, Chejanovsky N. Baculovirus mediated expression of a scorpion depressant toxin improves the insecticidal efficacy achieved with excitatory toxin. FEZiS Lett 1998; 422 : 132-6.

23. Fuxa JR, Richter AR, Ameen AO, Hammock BD. Vertical transmission of TnSNPV, TnCPV, AcMNPV and probably recombinant NPV in Trichoplusia ni. J Invertebr Pathol 2002; 79 : 44-50.

24. O' Brochta DA, Atkinson PW. Building the better bug. Sci Am 1998; 279 : 60-5.

25. Grace TDC. Establishment of a line of mosquito (Aedes aegypti, L) cells grown in vitro. Nature 1966; 211 : 366-7.

26. Greene AE, Charney J, Nickols WW, Corriel L. Species identity of insect cell lines. In Vitro 1972; 7 : 313-22.

27. Singh KRP. Cell cultures derived from larvae of Aedes albopictus (Skuse) and Aedes aegypti (L). Curr Sci 1967; 36 : 506-8.

28. Mitsuhashi J, Maramorosch K. Leafhopper tissue culture: embryonic, nymphal and imaginai tissues from aseptic insects. Contrib Boyce Thompson Inst 1964; 22 : 435-60.

29. Bhat UKM, Singh KRP. Establishment of a diploid cell line from larval tissues of Aedes vittatus (Bigot, 1861). Curr Sci 1970; 39 : 388-90.

30. Singh KRP, Bhat UKM. Establishment of two mosquito celllines from larval tissues of Aedes W-albus seperatum. Experientia 1971; 27 : 142-3.

31. Bhat UKM, Guru PY. Aedes novalbopictus: Establishment and characterization of larval cell lines. Exptl Parasitai 1973; 33 : 105-13.

32. Pant U, Dhanda V. Establishment of a cell line from Culex bitaeniorhynchus. J Tissue Culture Methods 1982; 6 : 61-3.

33. Pant U, Sudeep AB, Dhanda V, Mourya DT, Banerjee K. New embryonic cell line from Aedes krombeini (H) (Diptera: Culicidae). In Vitro Cell Dev BM (Animal) 1992; 28 : 567-8.

34. Athawale SS, Sudeep AB, Pant U, Berde PV, Jadi R, Mishra AC, et al. A new embryonic cell line from the embryonic tissues of Culex tritaeniorhynchus and its susceptibility to certain flaviviruses. Acta Virol 2002; 46 : 237-40.

35. Dhanda V, Kaul HN. Mosquito vectors of Japanese encephalitis virus and their bionomics in India. Proc Indian Natl Sci Acad 1980; B46 : 759-68.

36. Reuben R. Studies on the mosquitoes of North Arcot district, Madras state, India. Part I: Seasonal densities. J Med Entomol 1971; 8: 119-26.

37. Banerjee K, Deshmukh PK, Ilkal MA, Dhanda V. Transmission of JE virus by Culex bitaeniorhynchus (Giles). Indian J Med Res 1978; 67 : 889-93.

38. Pant U, Dhanda V. Arthropod tissue culture and its uses in arboviral research. ICMR Bull 1985; 16 : 33-8.

39. Pant U, Banerjee K, Athawale SS, Dhanda V. Susceptibility of Culex bitaeniorhynchus cell line to some arboviruses. Indian J Med Res 1985; 76 : 789-94.

40. Ghosh SN, Sathe PS, Sarathi SA, Cecilia D, Dandawate CN, Athawale SS, et al, Epitope analysis of strains of Japanese encephalitis virus by monoclonal antibodies. Indian J Med Res 1989; 89 : 368-75.

41. Pant U, Athawale SS, Sudeep AB. Susceptibility of Aedes krombeini cell line to some arboviruses. Indian J Med Res 1992; 95 : 239-44.

42. Sudeep AB, Pant U, Dhanda V, Banerjee K. A new Aedes krombeini cell line and its susceptibility to some arboviruses. In: Maramorosch, K. Mitsuhashi J, editors. Invertebrate cell culture: Novel directions and biotechnology applications. USA: Science Publishers Inc; 1997 p. 11-7.

43. Sudeep AB, Pant U. Aedes krombeini (H) cell line-A new approach to arboviral research. Paper presented at the IVth international symposium on vectors and vector borne diseases. Organized by the National Academy of Vector Borne Diseases, Bhubaneswar and Defence Research and Development Establishment, Gwalior, held at Gwalior, India from 18-20 February, 1999 p. 45.

44. Sreenivasan MA, Pant U. Athawale SS, Arankalle V. Detection by electron microscopy of endogenous viruses in Aedes krombeini (H.) line. In Vitro Cell Dev Biol (Animal) 1994; 3OA : 142-4.

45. Reuben R. A re-description of Culex vishnui (Theo) with notes on Culex pseudovishnui (Colles) and Culex tritaeniorhynchus (Giles) from southern India. Bull Entomol Res 1969; 58 : 643-52.

46. Singh KRP. Growth of arboviruses in arthropod tissue culture. Adv Virus Res 1972; 17 : 187-206.

47. Buckley, SM. Singh's Aedes albopictus cell cultures as helper cells for adaptation of Obodhiang and Kotonkhan viruses of the rabies sero-group to some vertebrate cell cultures. Appl Microbiol 1973; 25 : 695-6.

48. Singh KRP, Paul SD. Isolation of dengue virus in Aedes albopictus cell cultures. Bull World Health Organ 1969; 40 : 982-3.

49. Paul SD, Singh KRP. Comparative sensitivity of mosquito cell lines, VERO cell line and infant mouse with arboviruses. Curr Sd 1969; 10 : 241-2.

50. Chappel WA, Calisher CH, Toole RE, Maness KC, Sasso, DR, Henderson BE. Comparison of three methods used to isolate dengue virus type-2. Appl Microbiol 1971; 22 : 1100-3.

51. Gubler DJ, Kuno G, Sather GE, Valez M, Oliver A. Mosquito cell cultures and specific monoclonal antibodies in surveillance for DEN viruses. Am J Trop Med Hyg 1984; 33 : 158-65.

52. Ilkal MA, Dhanda V, Rodrigues JJ, Mohan Rao CVR, Moury a DT. Xenodiagnosis of laboratory acquired dengue infection by mosquito inoculation and immunofluorescence. Indian J Med Res 1984; 79 : 303-6.

53. Dhanda V, Mourya DT, Mishra AC, Ilkal MA, Pant U, Jacob PG, et al. Japanese encephalitis virus infection in mosquitoes reared from field collected immatures and in wild caught males. Am J Trop Med Hyg 1989; 41 : 732-6.

54. Mourya DT, Ilkal MA, Mishra AC, George Jacob P, Pant U, Ramanujam S, et al. Isolation of Japanese encephalitis virus from mosquitoes collected in Karnataka state, India from 1985-1987. Trans R Soc Trop Med Hyg 1989; 83 : 550-2.

55. Naik PS, Ilkal MA, Pant U, Kulkarni SM, Dhanda V. Isolation of Japanese encephalitis virus from Culex pseudovishnui Colless, 1957 (Diptera: Culicidae) in Goa. Indian J Med Res 1990; 91 : 331-3.

56. Pant U, Ilkal MA, Soman RS, Shetty PS, Kanojia PC, Kaul HN. First isolation of Japanese encephalitis virus from the mosquito Culex tritaeniorhynchus Giles, 1901 (Diptera: Culicidae) in Gorakhpur district, Uttar Pradesh. Indian J Med Res 1994; 99 : 149-51.

57. Igarashi A. Isolation of a Singh's Aedes albopictus cell clone sensitive to dengue and chikungunya viruses. J Gen Viral 1978; 40 : 531-44.

58. Varma MGR, Pudney M, Leake CJ. Cell lines from larvae of Aedes (Stegomyia) malayensis Colless and Aedes pseudoscutellaris (Theobald) and their infection with some arboviruses. Trans R Soc Trop Med Hyg 1974; 68: 374-82.

59. Yunker CE, Cory J. Plaque production by Singh's Aedes albopictus cells. Appl Microbiol 1975; 29 : 81-9.

60. Ajello C, Gresikova M, Buckley SM, Casals J. Detection of West Nile complement fixing antigens in Aedes albopictus cell cultures. Acta Virol 1975; 19 : 441-2.

61. Buckley, SM, Hayes CG, Maloney JM Lipman M, Aitken THG, Casals J. Arbovirus studies in invertebrate cell lines. In: Karstak E, Maramorosch K, editors. Invertebrate tissue culture, applications in medicine, biology and agriculture. New York: Academic Press; 1976; p. 3-19.

62. Soe thein, Auwanich W, Quina MA, Igarashi A, Okuno Y, Fukai K. Haemaglutinin prepared from cells of Ae. albopictus clone C6/36, infected with Type -1 dengue virus. Biken J 1979; 22 : 47-53.

63. Igarashi A, Morita K, Bundo K, Matsuo S, Hayashi K, Matsuo R, et al. Isolation of JE and GET viruses from Culex tritaeniorhynchus and slaughtered swine blood using Ae. albopictus clone C6 36 cells in Nagasaki, 1981. Trop Med 1981; 23 : 177-87.

64. Paul SD, Singh KRP, Bhat UKM. A study on the cytopathic effect of arboviruses on cultures from Aedes albopictus cell line. Indian J Med Res 1969; 57 : 339-48.

65. Pavri KM, Ghosh SN. Complement fixation tests for simultaneous isolation and identification of dengue virus using tissue culture. Bull World Health Organ 1969; 40 : 984-6.

66. Ghosh SN, Tongaokar SS, Dandavate CN. Intracellular and extracellular complement fixing antigens of JE and CHP viruses in mosquito tissue cultures. Curr Sci 1973; 42 : 286-8.

67. Ghosh SN, Bhat UKM. Hemagglutinating activity in normal culture fluid of Ae. albopictus cells. Curr Sci 1971; 40 : 354-5.

68. Padbidri VS, Pant U, Kulkarni SS, Dhanda V. Multiplication of Coxiella burnettii in Culex taeniorhynchus cell line. III International symposium on Rickettsia and Rickettsial Diseases, Smolence, Czechoslovakia, 1984; September 10-14.

69. Pant U, Banerjee K. Arthropod tissue culture in India (1967-1988)-A review. Indian J Med Res 1990; 91 : 397-407.

70. Kurti TJ, Munderloh UG. Mosquito tissue culture. Adv Cell Cult 1984; 3 : 259-302.

71. Fallen AM, Stollar V. The biochemistry and genetics of mosquito cells in culture. Adv Tissue Cult 1987; 5 : 97-137.

72. Fallen AM. Ribosome metabolism in mosquito cells. In: Kuroda Y, Kurstak E, Maramorosch K. editors. Invertebrate and fish tissue culture. Springer-Verlag: Japan Scientific Societies Press; 1988 p. 50-2.

73. Fallon AM, Sun D. Exploration of mosquito immunity using cells in culture. Insect Biochem Mol Biol 2001; 31 : 263-78.

74. Vlak JM, Hong HZ. Biosafety of engineered baculoviruses and cultured insect cells. In: Maramorosch K, Mitsuhashi J, editors. Invertebrate cell culture: Novel directions and biotechnology applications. USA: Science Publishers Inc; 1997 p. 181-91.

75. Heinz, KM, McCutchen, BF, Herrmann, R, Parella, MP, Hammock, BD. Direct effects of recombinant nuclear polyhedrosis viruses on selected non-target organisms. J Econ Entomol 1995; 88 : 259-64.

76. Luckow VA. Cloning and expression of heterologous genes in insect cells with baculovirus vectors. In: Prokop A, Bajpai RK, Ho CS, editors. Recombinant DNA technology and applications. New York: McGraw-Hill, Inc; 1991 p. 97-152.

77. Maeda S. Gene transfer vectors of a baculovirus, Bombyx mori nuclear polyhedrosis virus, and their use for expression of foreign genes in insect cells, In: Mitsuhashi, J, editor. Invertebrate cell system application. Boca Raton: CRC Press; 1989 p. 167-82.

78. Pant U, Mascarenhas AF, Jagannathan V. In vitro cultivation of a cell line from embryonic tissue of potato tuber moth, Gnorimoschema operculella (Zeller). Indian J Expl Biol 1977; 15 : 244-5.

79. Pant U, Athawale SS, Sudeep AB, Banerjee K. A new cell line from larval ovaries of Spodoptera litura (F) (Lepidoptera: Noctuidae). In Vitro Cell Dev Biol (Animal) 1997; 33 : 161-3

80. Pant U, Athawale SS, Basu A, Banerjee K. A new cell line established from pupal ovaries of Spodoptera litura (F) (Lepidoptera: Noctuidae). Indian J Exptl Biol 1998; 36 : 195-8.

81. Pant U, Athawale SS, Vipat VC. A new continuous cell line from larval hemocytes of Spodoptera litura (F). Indian J Exptl Biol 2000; 38 : 1201-6.

82. Sudeep AB, Shouche YS, Mourya DT, Pant U. New Helicoverpa armigera Hbn cell line from larval hemocytes for baculovirus studies. Indian J Exptl Biol 2002; 40 : 69-73.

83. Sudeep AB, Mourya DT, Shouche YS, Pidiyar V, Pant U. A new cell line from the embryonic tissue of Helicoverpa armigera Hbn (Lepidoptera: Noctuidae). In Vitro Cell Dev Biol (Animal) 2002; 38 : 262-4.

84. Sudeep AB, Mishra AC, Shouche YS, Pant U, Mourya DT. Establishment of two new cell lines from Bombyx mori (L) (Lepidoptera: Bombycidae) and their susceptibility to baculoviruses. Indian J Med Res 2002; 115 : 189-93.

85. Schneider I. Karyology of cell in culture, F. Characteristics of insect cells. In: Kruse PF Jr, Patterson MK Jr, editors. Tissue culture methods and applications. USA: Academic Press; 1973 p. 788-90.

86. Tabachnick WJ, Knudson DL. Characterization of invertebrate cell lines. II. Isoenzyme analyses employing starch gel electrophoresis. In Vitro 1980; 16 : 392-8.

87. Shouche YS, Patole MS, Pant U, Paranjpe S, Banerjee K. Authentication of two cell lines developed from the larval and pupal ovaries of Spodoptera litura by RNA based methods. In Vitro Cell Dev Biol (Animal) 1999; 35: 244-5.

88. Kshirasagar SG, Patole MS, Shouche YS. Characterization of insect cell lines: Heteroduplex analysis employing a mitochondrial 16S ribosomal RNA gene fragment. Anal Biochem 1997; 255 : 65-9.

89. Pant U, Sudeep AB, Athawale SS, Vipat VC. Baculovirus studies in new, indigenous lepidopteran cell lines. Indian J Exptl Biol 2002; 40 : 63-8.

90. Lua LHL, Reid S. Virus morphogenesis of Helicoverpa armigera nucleopolyhedrovirus in Helicoverpa zea serum free suspension culture. J Gen Virol 2000; 81 : 2531-43.

91. Sudeep AB. Establishment and characterization of new lepidopteran cell lines for baculovirus studies, Ph. D. Thesis. Pune: University of Pune; 2002.

92. Sriram S, Palhan VB, Gopinathan KP. Heterologous promoter recognition leading to high level of expression of cloned foreign genes in B. mori cell lines and larvae. Gene 1997; 161 : 181-9.

93. Saigal D, Gopinathan KP. Recombinant B. mori nucleopolyhedrovirus harboring green fluorescent protein. Biotechniques 1998; 25 : 997-1008.

94. Dalaya VP, Patil SP. Laboratory rearing and field release of Copidosoma kochleri Blanchard, and exotic parasite for control of Gnorimoschema operculella Zeller. Res J Mahatma Phule Agri Univ 1973; 4: 99-107.

95. Lery X, Giannotti J, Taha A, Ravale M, Abd-Ela S. Multiplication of a GV from PTM in a newly established cell line of Phthorimaea operculella. In Vitro Cell Dev Biol (Animal) 1997; 33 : 640-6.

96. Sudeep AB, Khushiramani R, Athawale SS, Mishra AC, Mourya DT. Characterization of a newly established potato tuber moth (Phthorimaea operculella Zeller) cell line. Indian J Med Res 2005; 121 : 159-63.

97. Palhan VB, Sumathy S, Gopinathan KP. Baculovirus mediated high-level expression of luciferase in silkworm cells and larvae. Biotechniques 1995; 19 : 97-104.

98. Bhat UKM, Singh KRP. In vitro culture of embryonic cells from and Indian triatomine bug, Triatoma rubrofasciatus De Geer (Hemiptera: Reduviidae). Curr Sci 1970; 39 : 488-9.

99. Pant U, Dhanda V, Argade SP, Banerjee K. Establishment of three cell lines from the embryonic tissue of Cimex hemipterus (F) (Hemiptera: Cimicidae). In Vitro Cell Dev BM (Animal) 1988; 24 : 1201-3.

100. Tamaz-Guerra P, McGuire MR, Behle RW, Shasha BS, Pingel RL. Storage stability of Anagrapha falcifera nucleopolyhedrovirus in spray-dried formulations. J Invertebr Pathol 2002; 79 : 7-16.

101. Luckow VA. Insect cell expression technology. In: Cleland JL, Craik CS, editors. Protein engineering: Principles and practice. New York: Wiley Liss Inc; 1996 p. 183-217.

A.B. Sudeep, D.T. Mourya & A.C. Mishra

National Institute of Virology (ICMR), Pune, India

Received March 25, 2003

Reprint requests: Dr A.B. Sudeep, Microbial Containment Complex (ICMR) Sus Road, Pushan, Pune 411021, India e-mail: sudeepnicc@yahoo.co.in

Copyright Indian Council of Medical Research Jun 2005
Provided by ProQuest Information and Learning Company. All rights Reserved

Return to Chikungunya
Home Contact Resources Exchange Links ebay