Biomedical subjects
A C Mishra
Publications and source records attributed to A C Mishra.
Venereal transmission of Chandipura virus by Phlebotomus papatasi (Scopoli).
Experiments were conducted in the laboratory on Phlebotomus papatasi to determine the possible role of males in maintaining or sustaining the Chandipura virus (CHPV) activity in nature. This study indicated that infected males are capable of passing on the virus to female sand flies while mating. The infection rate was found to be 12.5% in uninfected females when mated with infected males. The occurrence of venereal transmission of this virus may have epidemiologic importance in the natural cycle of CHPV.
Isolation of Ganjam virus from ticks collected off domestic animals around Pune, Maharashtra, India.
Studies on viruses of zoonotic importance in certain villages around Pune were undertaken between December 2000 and January 2002. A total of 1,138 adult ticks belonging to six different species were collected off domestic animals and processed for virus isolation. Six virus isolates were obtained. All six isolates were identified as Ganjam virus by Quick Complement Fixation test and reverse transcriptase-polymerase chain reaction using RNA nucleocapsid gene amplification. Five isolates were from the pools of adult Hemaphysalis intermedia ticks, and one isolate was from a pool of adult Rhipecephalus hemaphysaloides. This is the first report of isolation of Ganjam virus from Maharashtra state of India.
Detection of chandipura virus from sand flies in the genus Sergentomyia (Diptera: Phlebotomidae) at Karimnagar District, Andhra Pradesh, India.
A total of 191 adult sand flies belonging to the genus Sergentomyia were collected from seven villages in Karimnagar and Warangal districts of Andhra Pradesh State, India, after an outbreak of encephalitis due to Chandipura virus (CHPV). Fifteen pools, each containing two specimens, were tested by reverse transcriptase-polymerase chain reaction and sequencing. One pool of Sergentomyia from Kolanur village in Karimnagar District was positive for CHPV.
Vertical and venereal transmission of Chandipura virus (Rhabdoviridae) by Aedes aegypti (Diptera: Culicidae).
Experiments in the laboratory documented vertical and venereal transmission of Chandipura virus (CHPV) in Aedes aegypti (L.). The minimum filial infection rate among the progeny of infected females was 1.2%; the rate among male and female progeny was 0.9 and 1.4%, respectively. The venereal infection rate of CHPV among inseminated females was 32.7%. Our study indicates the possible occurrence of vertical and venereal transmission of CHPV in insect vectors.
Characterization of a newly established potato tuber moth (Phthorimaea operculella Zeller) cell line.
BACKGROUND & OBJECTIVE: Potato tuber moth (PTM), Phthorimaea operculella Zeller is a widely distributed, devastating pest of potatoes attacking the foliage and infest the tubers in both field and store causing serious economic damage. As application of PTM granulovirus (PTM-GV) has shown significant reduction in damage, attempts were made to develop a new cell line from this insect to grow PTM-GV for use as a biopesticide. METHODS: Approximately 100 mg of insect eggs were collected, surface sterilized and crushed gently in a boiling tube aseptically. The tissues were washed with physiological saline, suspended in growth medium and incubated stationary at 28 degrees C. Morphology of cells was studied after staining with Giemsa. Besides karyological and growth curve studies, PCR amplification was also done for rapid amplified polymorphic DNA pattern. RESULTS: A new cell line from the embryonic tissue of PTM was maintained in Mitsuhashi Maramorosch medium supplemented with 10 per cent foetal bovine serum. It is in the 78th passage level and designated as NIV-PTM-1095. Random amplified polymorphic DNA profile analysis indicated this as a new cell line from potato tuber moth and differed from the profiles of two other lepidopteran cell lines maintained in the laboratory. Three different cell types were observed at the 40th passage level and comprised of epithelial-like cells (77%), fibroblast-like cells (20%) and giant cells (3%). The chromosome number varied from 54-176. The cell line had a cell doubling time of approximately 42 h during the logarithmic phase of growth. The cell line did not support the multiplication of any of the baculoviruses used in the study. INTERPRETATION & CONCLUSION: Since the new cell line is found to replicate PTM-GV, it may be useful for the propagation of PTM-GV in large scale. Studies to scale up the production of the GV in the cell line and field trials may lead to its widespread use as an eco-friendly biopesticide.
Incidence of neutralizing antibodies to Chandipura virus in domestic animals from Karimnagar and Warangal Districts of Andhra Pradesh, India.
In order to determine the possible role of domestic animals in the outbreak of acute encephalitis associated with Chandipura virus (CPV) among children in Andhra Pradesh in 2003, a serological survey of domestic animals was carried out during the epidemic in July 2003. Out of 180 animal sera from highly affected areas of the Karimnagar and Warangal districts of Andhra Pradesh 33 (18.3%) had virus neutralizing (VN) antibodies to CPV. The positive animals consisted of pigs (30.6%), buffalos (17.9%), cattle (14.3%), goats (9.3%) and sheep (7.7%). Isolation of CPV and detection of CPV antibodies in patients with encephalitis reported earlier and the evidence of antibodies to CPV in domestic animals shown here suggest that CPV circulates in this region and should be considered an emerging virus of public health importance.
Insect cell culture in research: Indian scenario.
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.
An outbreak of Chandipura virus encephalitis in the eastern districts of Gujarat state, India.
An outbreak of encephalitis with a case fatality rate of 78.3% was investigated among children in Gujarat State, India. Twenty-six cases were reported. Three patients had IgM antibodies to Chandipura virus. Virus was isolated from one patient with rhabdomyosarcoma in porcine stable cell lines and in suckling mice. Chandipura virus RNA was present in 9 of 20 acute-phase serum samples, and virus sequences from the present outbreak were closely related to prototype strain (1965) and Andhra Pradesh, India (2003) isolates. Serologic and molecular assays documented the absence of Japanese encephalitis virus, West Nile virus, dengue virus, and paramyxoviruses in clinical samples. The etiologic agent was Chandipura virus, which has become an important encephalitis-causing virus in India.
Effect of temperature and insecticide stresses on Aedes aegypti larvae and their influence on the susceptibility of mosquitoes to dengue-2 virus.
Two major factors, higher temperatures and the application of insecticides, can drastically alter the genetic structure of a vector mosquito population. Due to these two stresses, the majority of the population gets wiped out, but the ones that withstand the stress and survive are likely to pass on survivability, and have an altered physiology. Our study shows that exposures to higher temperatures and DDT during the larval stage affects their susceptibility as adult mosquitoes to the DEN-2 virus. The overall transcription and translation status of heat shock protein (Hsp70) in virus high- and low-susceptible was the same as that in other batches. In the case of a DDT-resistant (R-7) strain two bands were obtained during RT-PCRs after heat shock. These two alleles were obtained only with HY-1 in which R-7 males were used for the crosses, suggesting that the second allele is probably male sex linked. The higher expression of Hsp70 may provide DDT-resistant strains a better chance of survival high temperature environments, particularly in homozygotes and hybrids. It was also interesting to note that these strains have a significantly lower susceptibility to the virus. Wide-spread DDT-resistance and a rise in temperature above the average temperature during summer may result in a population with a low susceptibility to the virus. Several families of heat shock proteins are known to be expressed in mosquitoes, and may have a cumulative role in determining susceptibility to the virus, which itself is governed by several genes.
Effect of temperature stress on immature stages and susceptibility of Aedes aegypti mosquitoes to chikungunya virus.
A high temperature stress of 44.5 degrees C for 10 minutes on the larval stages was found to affect the susceptibility of adult Aedes aegypti mosquitoes to chikungunya virus. At this temperature, the mortality of the mosquito larvae was found to be approximately 95%, whereas a temperature greater than 45 degrees C for 10 minutes was found to be lethal. A temperature tolerant (TT) strain was developed by exposing the larvae to a temperature of 44.5 degrees C for 10 minutes at every generation for five generations. This strain was established to determine whether increase in the susceptibility was due to any selection pressure of higher temperature or to the influence of other intrinsic factors such as expression of immunoresponsive (IR) genes. Other studies on these mosquito strains showed that when maintained at 28 +/- 1 degrees C, there was no difference in the larval duration and mortality in the immature stages, but the mean survival of female mosquitoes in the TT strain was 5-6 days longer. Conversely, when mosquitoes were maintained throughout at 37 degrees C the mean survival of the mosquitoes decreased drastically in both strains, but the mean survival of females in the TT strain was 5-6 days longer compared with the unstressed controls. This increases the probability of at least one more blood meal. Fecundity of the TT strain was found to be lower than that of the control mosquitoes. Data suggest that expression of certain IR genes was affected by the heat shock. Some of these genes were up-regulated and down-regulated, which may have affected the susceptibility of mosquitoes to the virus. Although there was some selection in the temperature-tolerant individuals in the TT strain, when stressed by heat they showed expression of IR genes in a pattern similar to that in the normal controls. It appears that an increase in temperature above the average temperature of an area might help increase the proportion of virus-susceptible mosquitoes in the population. Such an increase in temperature in an endemic area would not only enhance the selection of temperature-tolerant individuals in a population having more longevity, but would also affect both intrinsic and extrinsic factors by reducing the extrinsic incubation period and increasing susceptibility of mosquitoes to viruses due to affected expression of IR genes.
The role of hemagglutinins in the midgut extracts of two lines of Aedes aegypti in their susceptibility to Dengue-2 virus.
Hemagglutinin activity (HA) was studied in the midgut extracts from highly (h) and lowly susceptible strains of Aedes aegypti mosquitoes to Dengue-2 virus (DEN-2). HA in the midgut extracts from these two isofemale strains of mosquitoes was high in as compared to (h) mosquitoes. HA was found to be higher with chicken red blood cells (RBCs) than with rabbit and human RBCs of O group. Larval midgut extracts showed higher activity than those from adult female mosquitoes. Exposure of midgut extracts to 100 degrees C for 10 mins destroyed the activity. The activity was observed between pH 6 and pH 10. HA in midgut extracts was also studied using twenty different carbohydrates; five of them showed an inhibition of HA. The inhibitory carbohydrates, when incorporated into DEN-2-infected bloodmeal, showed a reduction in the susceptibility of mosquitoes to the virus as compared to the control ones fed on the virus alone. Similarly, when these carbohydrates were incorporated in the DEN-2-infected inoculum, the inoculated mosquitoes showed a reduction in the susceptibility to the virus. HA in the virus-infected midgut extracts was higher than that in the uninfected controls. These results suggest that the presence of HA in the midgut may be one of the factors that affect the susceptibility of Ae. aegypti mosquitoes to DEN-2.
A preliminary study of multilevel geographic distribution & prevalence of Aedes aegypti (Diptera: Culicidae) in the state of Goa, India.
BACKGROUND & OBJECTIVES: Dengue virus activity has never been reported in the state of Goa. The present study was carried out to document a multilevel geographic distribution, prevalence and preliminary analysis of risk factors for the invasions of Aedes aegypti in Goa. METHODS: A geographic information system (GIS) based Ae. aegypti surveys were conducted in dry (April 2002) and wet (July 2002) seasons in the rural and urban settlements. The random walk method was used for household coverage. The non-residential area visits included ancillaries of roadways, railways, air-and seaports. Simultaneous adult mosquito collections and one-larva per container technique were adopted. RESULTS: The Ae. aegypti larval and adult prevalence was noted in all the four urban areas in both dry (Density index (DI)= 3 to 6) and wet (DI= 5 to 7) seasons and only one out of 3 villages showed Ae aegypti presence in wet season (DI= 5 to 7). In the residential areas, hutments showed higher relative prevalence indices (Breteau index, BI=100; container index, CI=11.95; adult house index, AHI=13.33) followed by close set cement houses (BI=44.1; CI=12.0; AHI=11.24). Ae aegypti relative prevalence indices were also more for households with pets (BI=85.11; CI=12.5; AHI= 42.85); those with tap had higher risk (larval house index, LHI =32.03; relative risk, RR>2, n=256). Plastic drum was the most preferred breeding place (chi(2) = 19.81; P<0.01; RR=3.41) among domestic containers and rubber tyres (chi(2) = 11.86; P<0.01; RR=3.61)among sundry/rainfilled containers. INTERPRETATION & CONCLUSION: Established Ae aegypti prevalence in the urban settlements during dry and wet seasons and its scattered distribution in a rural settlement spell risk of dengue infection at macro-level. In the residential areas nature and types of the households, tap water supply and storage and communities' attitude and practices contribute to sustained meso-level risk of Ae aegypti prevalence dependant DEN. The non-residential areas offer transient meso-level risk as Ae aegypti prevalence was seasonally unstable and monsoon dependent. Risk at micro-level was due to the preferred larval habitats of Ae aegypti breeding viz., residential plastic-ware and tyres, and transport tyres in non-residential areas.
Experimental transmission of Chikungunya virus by Anopheles stephensi mosquitoes.
The Ades aegypti mosquito has been considered the principal vector of Chikungunya (CHIK) virus. As CHIK epidemics usually occur in urban regions and Anopheles stephensi is another highly endophilic and anthropophilic mosquito, there is a very high probability of this mosquito to feed on CHIK virus-infected patients, to pick up and transmit the virus. Therefore the present study was conducted to test the CHIK virus transmission capability for the A. stephensi mosquito. The obtained results showed that this mosquito species is capable of transmitting CHIK virus. It is surmised that during any epidemic of febrile illness CHIK virus isolation attempts should also be made from this mosquito species.
Genotyping of Chikungunya virus isolates from India during 1963-2000 by reverse transcription-polymerase chain reaction.
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Serological evidence for Japanese encephalitis virus and West Nile virus infections in water frequenting and terrestrial wild birds in Kolar District, Karnataka State, India. A retrospective study.
In a serological survey of birds in a Japanese encephalitis (JE) endemic area of Kolar District, Karnataka State, India, 859 bird sera were tested by hemagglutination-inhibition test (HIT) for JE encephalitis and West Nile encephalitis (WNE) viruses. Only 2 (0.002%) and 178 (20.72%) sera were positive for JE virus (JEV) and WNE virus (WNV), respectively. Only 160 (18.63%) of 859 sera could be subjected to neutralizing test (NT). Of these, 20 (12.50%) and 62 (38.75%) were positive for JEV and WNV antibodies, respectively. These findings indicate that bird species such as Pond Herons and Little Egrets among ardeid birds and Grey Partridges and Quails among terrestrial birds are infected with JEV and WNV and play probably a role in the maintenance of these viruses in the abovementioned part of India.
West Nile virus: the Indian scenario.
West Nile virus (WNV) is an important arthropod borne flavivirus; usually causes a mild infection called West Nile fever (WNF) in human and horses. Mosquitoes are the principal vectors of WNV. Various Culex species are found to act as vectors in different geographical regions. The virus is maintained in a bird-mosquito cycle in nature. In India, Culex mosquitoes are tentatively incriminated as vectors of WNV. Experimental studies have shown that Culex tritaeniorhynchus, Cx. vishnui, Cx. bitaeniorhynchus and Cx. univittatus, Culex pipiens fatigans and Aedes albopictus could act as potential vectors of WNV. Transovarial transmission of WNV has been experimentally demonstrated in Culex mosquitoes. Apart from mosquitoes, the role of other arthropods is also considered in the maintenance of WNV during inter-enzootic periods. The possible role of ardeid birds in the maintenance of WNV has been described in India. Though very few clinically overt cases of human encephalitis due to WNV are observed, Japanese encephalitis virus (JEV) is found to dominate in southern India. WNF in horses has not been documented in India. JEV immunized monkeys were protected from WNV challenge and the WNV immunization was found to reduce the disease severity due to JEV. Based on the limited genome sequence analysis, the Indian isolates are grouped together under the genetic lineage-I. WNV infection is diagnosed by IgM antibody capture enzyme linked immunosorbant assay, haemagglutination inhibition test, neutralization test and reverse transcriptase-polymerase chain reaction (RT-PCR). For the effective control of Culex mosquitoes, integrated vector control strategies are recommended. Specific methods are not available for the treatment of WNV infection. However, in patients with encephalitis supportive therapy is recommended. Though a few candidate vaccines are under laboratory trial, no vaccine has been available commercially for the control of WNV infection in human and animals. In view of the global interest on WNV, this paper describes the present status of WNV in India.
A focal outbreak of Japanese Encephalitis among horses in Pune district, India.
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