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'Candidatus Liberibacter asiaticus' (CLas) is a non-culturable α-proteobacterium, putatively causing Huanglongbing (HLB), a devastating disease currently threatening citrus production in India and all over the world. Here, we report the whole genome sequence of a CLas strain obtained from the leaf midrib of a HLB-infected Khasi mandarin tree in Assam, India. The draft genome of strain AS-TNSK3 is 1236955 bp with G+C content of 36.5% and harbours both Type 1- and Type 2- like prophages. Annotation of AS-TNSK3 genome identifies 1084 open reading frames (ORFs) and 45 RNA genes. This CLas whole genome sequence is the first report from Northeast India, considered to be the centre of origin of the genus Citrus. Insights from the Indian CLas sequence data would enrich the current CLas genome resources and will facilitate in understanding CLas biology and designing effective HLB disease management strategy. [ABSTRACT FROM AUTHOR]

The Asian citrus psyllid, Diaphorina citri Kuwayama (Hemiptera: Psyllidae) is the vector of Candidatus Liberibacter asiaticus (CLas), a proteobacterium associated with the most lethal Huanglongbing (HLB) or citrus greening disease. This study was aimed to detect CLas in naturally occurring D. citri vectors and to find out the genetic interconnection and diversity among the populations of D. citri and CLas in the citrus belts of India. Morphological and molecular identification confirmed the 561 adult psyllid samples, collected from 32 citrus orchards located in 8 states of India, as D. citri. Sequence data of two different genes, cytochrome oxidase subunit I (mtCOI), and 16S rRNA, were used to characterize the populations of D. citri and CLas, respectively. In the mtCOI-based phylogenetic tree, two major genetic clusters were observed (A and B) and further in the cluster B, four genetic subgroups (B1, B2, B3 and B4) were noticed. The tree topology clearly indicated that D. citri mtCOI sequences obtained from eight different Indian states separated into four different clades: A, B1, B2 and B4, composed of mtCOI haplotypes Dcit-6, Dcit-2, Dcit-23 and Dcit-1, respectively. Presence of CLas was also validated in the psyllid vector through quantitative real time PCR, conventional PCR and phylogenetic analysis of 16S rRNA gene sequences. These findings may be helpful in the development of region-wide management strategy to be adopted for controlling the D. citri and/or the HLB disease it vectors in different citrus production zones of India. [ABSTRACT FROM AUTHOR]

Huanglongbing (HLB), also known as 'citrus greening', is an extremely destructive disease of citrus worldwide. HLB is associated with three species of the fastidious proteobacterium, Candidatus Liberibacter asiaticus (CaLas), Ca. L. africanus and Ca. L. americanus with CaLas being the most widely distributed around the world and the only species detected and described so far in India, one of the major global citrus fruit producers. Prophages are highly dynamic components in the bacterial genome and play an important role in intraspecies variations. Three types of prophages, Type 1, Type 2 and Type 3 have been identified and described in CaLas so far. In the present study, 441 CaLas isolates sampled across 18 Indian states were used for prophage typing. Based on detection of three prophage types by PCR, all the eight probable combinations of CaLas prophages were identified, including single Type 1 (26.5%), single Type 2 (18.8%), single Type 3 (1.4%), Type 1 + Type 2 (20.4%), Type 1 + Type 3 (12.5%), Type 2 + Type 3 (4.8%), Type 1 + Type 2 + Type 3 (11.3%) and None type (4.3%). Prophage types were confirmed by PCR amplicon sequencing and subsequent phylogenetic analysis. By discovery of all 3 prophages and based on genetic identity and genetic distance, CaLas populations from eighteen citrus growing states were separated into two major Prophage Typing Groups (PTGs): PTG1 and PTG2. The PTG1 comprised of CaLas from North-West India and PTG2 from rest of the country (North-East, Central and South India), and both major groups were further divided into two (PTG1-A, PTG1-B) and three (PTG2-A, PTG2-B and PTG2-C) subgroups respectively. The findings of CaLas population patterns provide evidence for independent origins of HLB-associated CaLas. CRISPR (clustered regularly interspaced short palindromic repeats) array was also detected in CaLas isolates. This is the first report evaluating the genetic variation of a large population of CaLas bacterium in India using the PCR markers from the prophage regions which would certainly assist the ongoing HLB management efforts in India. [ABSTRACT FROM AUTHOR]

Kinnow mandarin is an important commercial fruit crop in India. APhytophthoraspecies was recovered from rhizosphere soil of declining Kinnow mandarin trees at Abohar area, Punjab state, India in January 2015. Morphological and physiological characteristics coupled with ITS-RFLP and sequence analysis of the ITS andcox1 gene regions revealed the isolate was an atypical strain ofP. inundata. The isolate had ovoid to limoniform non-papillate sporangia and produced thick-walled, spherical oogonia with amphigynous antheridia. Thin-walled chlamydospores or hyphal swellings with radiating hyphae were formed readily in water in greater abundance than sporangia. This atypical strain ofP. inundatawas heterothallic (A1 mating type) and grew very slowly at 35°C. The optimum temperature for growth was 25°C and the isolate was metalaxyl-sensitive. Pathogenicity tests indicated that the strain ofP. inundatahas the potential to infect leaves and stems of Kinnow mandarin and was also pathogenic to rough lemon rootstock seedlings, causing root rot and decline. To our knowledge, this is the first report ofP. inundatain India. [ABSTRACT FROM PUBLISHER]

The widespread occurrence of Huanglongbing ( HLB) was recorded in sixteen citrus growing states of India using the real-time quantitative PCR and the derived threshold cycle ( Ct) value. All the commercially important citrus varieties of mandarin, sweet orange, lime and lemon, pummelo and Satkara were infected with ' Candidatus Liberibacter asiaticus', the bacterium associated with HLB. Ct values positive for HLB were found in all the states except Arunachal Pradesh. The primer-probe combination HLBas- HLBr- HLBp was found specific to Ca. L. asiaticus and do not exhibit any cross-reactivity with other pathogenic residents of citrus. [ABSTRACT FROM AUTHOR]

Background: The focus of India's National Malaria Programme witnessed a paradigm shift recently from health facility to community-based approaches. The current thrust is on diagnosing and treating malaria by community health workers and prevention through free provision of long-lasting insecticidal nets. However, appropriate community awareness and practice are inevitable for the effectiveness of such efforts. In this context, the study assessed community perceptions and practice on malaria and similar febrile illnesses. This evidence base is intended to direct the roll-out of the new strategies and improve community acceptance and utilization of services. Methods: A qualitative study involving 26 focus group discussions and 40 key informant interviews was conducted in two districts of Odisha State in India. The key points of discussion were centred on community perceptions and practice regarding malaria prevention and treatment. Thematic analysis of data was performed. Results: The 272 respondents consisted of 50% females, three-quarter scheduled tribe community and 30% students. A half of them were literates. Malaria was reported to be the most common disease in their settings with multiple modes of transmission by the FGD participants. Adoption of prevention methods was seasonal with perceived mosquito density. The reported use of bed nets was low and the utilization was determined by seasonality, affordability, intoxication and alternate uses of nets. Although respondents were aware of malaria-related symptoms, care-seeking from traditional healers and unqualified providers was prevalent. The respondents expressed lack of trust in the community health workers due to frequent drug stock-outs. The major determinants of health care seeking were socio-cultural beliefs, age, gender, faith in the service provider, proximity, poverty, and perceived effectiveness of available services. Conclusion: Apart from the socio-cultural and behavioural factors, the availability of acceptable care can modulate the community perceptions and practices on malaria management. The current community awareness on symptoms of malaria and prevention is fair, yet the prevention and treatment practices are not optimal. Promoting active community involvement and ownership in malaria control and management through strengthening community based organizations would be relevant. Further, timely availability of drugs and commodities at the community level can improve their confidence in the public health system. [ABSTRACT FROM AUTHOR]

Objective: India reported the highest number of road traffic crashes, related injuries, and deaths among all countries in the world, with 105,725 road traffic fatalities and 452,922 nonfatal road traffic injuries in 2007. In this report we present a systematic review of available literature on the use of psychoactive substances (alcohol and drugs) among road users, particularly those involved in road traffic crashes (RTCs). Methods: MEDLINE, EMBASE, Ind Medica, and several other databases were searched for reports published between 1980 and 2011 that present data on the prevalence or extent of substance use among road users in India. Results: Among the 23 studies eligible for the review, alcohol was reported by all, but only 2 mentioned the use of drugs. Most of the studies were hospital based, included injured or killed road users, and belonged to southern parts of India. Seven studies did not report any method for detecting alcohol use, whereas 7 used analytical testing, 7 used self-reporting, and 2 used observation. Utilizing the various means of verification, the studies reported that 2 to 33 percent of injured and 6 to 48 percent of killed RTC victims had consumed alcohol or drugs; only 2 mentioned drugs without specifying which types. Most studies did not distinguish between drivers, passengers, bicyclists, and pedestrians, and none investigated alcohol or drug use among those responsible for the accident. Conclusion: A significant proportion of injured or killed road users in India had used alcohol before the accident. However, the existing studies cannot be used to estimate the risk of accident involvement among drunk drivers. There is a need for more rigorous research and capacity building on substance use vis-à-vis road traffic crashes. [ABSTRACT FROM PUBLISHER]

Background: Orissa state in eastern India accounts for the highest malaria burden to the nation. However, evidences are limited on its treatment-seeking behaviour in the state. We assessed the treatment-seeking behaviour towards febrile illness in a malaria endemic district in Orissa. Methods: A cross-sectional community-based survey was carried out during the high malaria transmission season of 2006 in Boudh district. Respondents (n = 300) who had fever with chills within two weeks prior to the day of data collection were selected through a multi-stage sampling and interviewed with a pre-tested and structured interview schedule. Malaria treatment providers (n = 23) were interviewed in the district to gather their insights on factors associated with prompt and effective treatment through a semi-structured and open-ended interview guideline. Results: Majority of respondents (n = 281) sought some sort of treatment e.g. government health facility (35.7%), less qualified providers (31.3%), and community level health workers and volunteers (24.3%). The single most common reason (66.9%) for choosing a provider was proximity. Over a half (55.7%) sought treatment from appropriate providers within 48 hours of onset of symptoms. Respondents under five years (OR 2.00, 95% CI 0.84-4.80, P = 0.012), belonging to scheduled tribe community (OR 2.13, 95% CI 1.11-4.07, P = 0.022) and visiting a provider more than five kilometers (OR 2.04, 95% CI 1.09-3.83, P = 0.026) were more likely to have delayed or inappropriate treatment. Interviews with the providers indicated that patients' lack of trust in community volunteers providing treatment led to inappropriate treatment-seeking from the less qualified providers. The reasons for the lack of trust included drug side effects, suspicions about drug quality, stock-outs of drugs and inappropriate attitude of the provider. Conclusion: Large-scale involvement of less qualified providers is suggested in the malaria control programme as volunteers after appropriate capacity development since the community has more trust in them. This should be supported by uninterrupted supply of drugs to the community volunteers, and involvement of the community-based organizations and volunteers in the planning, implementation, and monitoring of malaria control services. There is also a need for continuous and rigorous impact evaluations of the program to make necessary modifications, scale up and to prevent drug resistance. [ABSTRACT FROM AUTHOR]

We report 11 cases of severe Plasmodium vivax malaria in Bikaner (western India). Patients exhibited cerebral malaria, renal failure, circulatory collapse, severe anemia, hemoglobinurea, abnormal bleeding, acute respiratory distress syndrome, and jaundice. Peripheral blood microscopy, parasite antigen-based assays, and parasite 18s rRNA gene-based polymerase chain reaction showed the presence of P, vivax and absence of P, falciparum,. [ABSTRACT FROM AUTHOR]

Abstract: Malaria parasites exhibit sequence diversity for a number of stage specific antigens. Several studies have proved that apical membrane antigen-1 (AMA-1) is an effective target for eliciting a protective immune response in humans and other experimental animals. We have investigated the sequence variation in Plasmodium vivax AMA-1 (Pv AMA-1) from different Indian isolates. This is the first study of its kind for the nearly full length Pv AMA-1 from India. Our analysis reveals greater degree of genetic diversity in Pv AMA-1 than reported so far and identifies five novel haplotypes. This is significant to establish the antigenic repertoire of isolates in a malaria endemic country like India. [Copyright &y& Elsevier]

Abstract: Malaria parasites exhibit sequence diversity for a number of stage specific antigens. Several studies have proved that merozoite surface protein-1 (MSP-1) is an effective target eliciting a protective immune response. The MSP-142 region comprising two EGF-like domains is involved in generating protective immune response in humans and other experimental animals. Searching for point mutations in this region is essential in view of vaccine development. We have investigated the sequence variations in Plasmodium falciparum MSP-1 carboxy terminal region in field isolates from different regions in India. Our study reveals the presence of eight variant types of MSP-119 in the Indian sub-continent, which comprise of E-TSR-L, Q-TSR-L, E-TSG-L, Q-KNG-L, Q-KNG-F, E-KNG-L, E-KNG-F, and E-KYG-F. The last named allele is a novel variant being reported for the first time. [Copyright &y& Elsevier]

Background: Malaria continues to be a prominent global public health challenge. This study tested the effectiveness of two service delivery models for reducing the malaria burden, e.g. supportive supervision of community health workers (CHW) and community mobilization in promoting appropriate health-seeking behaviour for febrile illnesses in Odisha, India., Methods: The study population comprised 120 villages from two purposively chosen malaria-endemic districts, with 40 villages randomly assigned to each of the two treatment arms, one with both supportive supervision and community mobilization and one with community mobilization alone, as well as an observational control arm. Outcome measures included changes in the utilization of bed nets and timely care-seeking for fever from a trained provider compared to the control group. Analysis was by intention-to-treat., Results: Significant improvements were observed in the reported utilization of bed nets in both intervention arms (84.5% in arm A and 82.4% in arm B versus 78.6% in the control arm; p < 0.001). While overall rates of treatment-seeking were equal across study arms, treatment-seeking from a CHW was higher in both intervention arms (28%; p = 0.005 and 27.6%; p = 0.007) than in the control arm (19.2%). Fever cases were significantly more likely to visit a CHW and receive a timely diagnosis of fever in the combined interventions arm than in the control arm (82.1% vs. 67.1%; p = 0.025). Care-seeking from trained providers also increased with a substitution away from untrained providers. Further, fever cases from the combined interventions arm (60.6%; p = 0.004) and the community mobilization arm (59.3%; p = 0.012) were more likely to have received treatment from a skilled provider within 24 hours than fever cases from the control arm (50.1%). In particular, women from the combined interventions arm were more likely to have received timely treatment from a skilled provider (61.6% vs. 47.2%; p = 0.028)., Conclusion: A community-based intervention combining the supportive supervision of community health workers with intensive community mobilization and can be effective in improving care-seeking and preventive behaviour and may be used to strengthen the national malaria control programme.

Background & Objectives: Description of severe vivax malaria and mixed species infection requires good clinical study. The present study was undertaken to evalute the characteristics of severe malaria patients in Bikaner, northwest India., Methods: This prospective study included 539 admitted adult patients of severe malaria (Plasmodium falciparum 274, P. vivax 221, and mixed infection of Pv + Pf 44). The diagnosis was confirmed by polymerase chain reaction. The categorization of severe malaria was done strictly as per WHO criteria., Results: The distribution of severe manifestation was similar in severe vivax, falciparum and mixed infections except more cases of thrombocytopenia in P. vivax (p=0.030) and in mixed infection (p=0.004). The risk of developing severe malaria was greatest in patients of mixed infection [53.01% (44/83)] in comparison to Plasmodium falciparum malaria [49.37% (274/555), RR= 1.135; p=0.616] and P. vivax malaria [45.38% (221/ 487), RR = 1.299, p=0.243]. Hepatic dysfunction was the commonest pernicious syndrome [P. falciparum 50% (137/274), P. vivax 43.89% (97/221), and mixed infections 54.55% (24/44)]. Multiorgan dysfunction was present in 40.26% (217/539) patients, the risk was greatest in mixed infection [90.90% (40/44)] in comparison to P. falciparum monoinfection [37.59% (103/274), RR = 12.238; p=0.0001] or P. vivax monoinfection [33.48% (74/ 221), RR = 13.25; p=0.0001]. The risk of mortality in severe malaria was 6.31% (34/539) in which mixed infection had greater risk [9.09% (4/44)] in comparison to P. falciparum [7.30% (20/274); OR = 1.270 (CI 0.347-4.217); p=0.757] or P. vivax [4.52% (10/221); 0R 2.110 (CI 0.527-7.826); p=0.260]., Interpretation & Conclusion: Severe vivax or falciparum malaria had almost similar features and prognosis including mortality. Risk of developing severe malaria, multiorgan dysfunction and mortality was more in patients of mixed infection in comparison to P. falciparum or P. vivax monoinfection. A multicentric study on larger number of patients requires further confirmation.

The 28S rRNA gene was amplified and sequenced from P. falciparum and P. vivax isolates collected from northwest India. Based upon the sequence diversity of the Plasmodium 28SrRNA gene in comparison with its human counterpart, various nested polymerase chain reaction (PCR) primers were designed from the 3R region of the 28SrRNA gene and evaluated on field isolates. This is the first report demonstrating the utility of this gene for species-specific diagnosis of malaria for these two species, prevalent in India. The initial evaluation on 363 clinical isolates indicated that, in comparison with microscopy, which showed sensitivity and specificity of 85·39% and 100% respectively, the sensitivity and specificity of the nested PCR assay was found to be 99·08% and 100% respectively. This assay was also successful in detecting mixed infections that are undetected by microscopy. Our results demonstrate the utility of the 28S rRNA gene as a diagnostic target for the detection of the major plasmodial species infecting humans.

Objective: To evaluate microscopy, OptiMAL(®) and multiplex PCR for the identification of Plasmodium falciparumm (P. falciparum) and Plasmodium vivax (P. vivax) from the field isolates of Bikaner, Rajasthan (Northwest India)., Methods: In this study, a multiplex PCR (P. falciparum and P. vivax) was further developed with the incorporation of Plasmodium malariae (P. malariae) specific primer and also a positive control. The performance of microscopy, plasmodium lactate dehydrogenase (pLDH) based malaria rapid diagnostic test OptiMAL(®) and 18S rRNA gene based multiplex PCR for the diagnosis of P. falciparum and P. vivax was compared., Results: The three species multiplex PCR (P. falciparum, P. vivax and P. malariae) with an inbuilt positive control was developed and evaluated. In comparison with multiplex PCR, which showed the sensitivity and specificity of 99.36% (95%CI, 98.11%-100.00%) and 100.00% (95%CI, 100.00%-100.00%), the sensitivity and specificity of microscopy was 90.44% (95%CI, 88.84%-95.04%) and 99.22% (95%CI, 97.71%-100.00%), and OptiMAL(®) was 93.58% (95%CI, 89.75%-97.42%) and 97.69% (95%CI, 95.10%-100.00%). The efficiencies were 99.65%, 95.10% and 95.45% for multiplex PCR, microscopy and OptiMAL(®), respectively., Conclusions: Our results raise concerns over the overall sensitivities of microscopy and OptiMAL(®), when compared to the multiplex PCR and thus stress the need for new molecular interventions in the accurate detection of the malarial parasites. This further highlights the fact that further developments are needed to improve the performance of rapid diagnostic tests at field level., (Copyright © 2013 Hainan Medical College. Published by Elsevier B.V. All rights reserved.)

Plasmodium vivax is the predominant species of the human malaria parasite present in the Indian subcontinent. There have been recent reports on Chloroquine (CQ) resistance and severe manifestations shown by P. vivax from different regions of the world including India. This study focuses on Bikaner, India where during the last few years there have been continuous reports of severe manifestations by both Plasmodium falciparum and P. vivax. This region has a widespread use of Chloroquine and Sulfadoxine-Pyrimethamine for the treatment of malaria, but the resistance profiles of these drugs are not available. We report here the profile of mutations in marker genes associated with Chloroquine and antifolate drug resistance among the P. vivax parasites obtained from patients with severe (n=30) and non-severe (n=48) manifestations from this region. Most isolates showed the wild type alleles for both the Chloroquine and antifolate resistance markers (P<0.0005). Except for one isolate showing Y976F mutation in the Pvmdr-1 gene, no reported mutation was observed in the Pvmdr-1 or Pvcrt gene. This is in accordance with the fact that till date no Chloroquine resistance has been reported from this region. However, the single isolate with a mutation in Pvmdr-1 may suggest the beginning of the trend towards decreased susceptibility to Chloroquine. The frequency of PvDHFR-PvDHPS two locus mutations was higher among the patients showing severe manifestations than the patient group with non-severe (uncomplicated) malaria (P<0.003). None of the parasites from patients with uncomplicated P. vivax malaria showed the mutant PvDHPS genotype. Novel mutations in PvDHFR (S117H) and PvDHPS (F365L, D459A and M601I) were observed only in the parasite population obtained from patients exhibiting severe complications. Preliminary homology modeling and molecular docking studies predicted that these mutations apparently do not have any effect on the binding of the drug molecule to the enzyme. However, the presence of novel mutations in the PvDHPS gene indicate a degree of polymorphism of this molecule which is in contrast to available published information., (Copyright © 2012 Elsevier Inc. All rights reserved.)

The apicomplexan parasite Plasmodium vivax is responsible for causing more than 70% of human malaria cases in Central and South America, Southeastern Asia and the Indian subcontinent. The rising severity of the disease and the increasing incidences of resistance shown by this parasite towards usual therapeutic regimens have necessitated investigation of putative novel drug targets to combat this disease. The apicoplast, an organelle of procaryotic origin, and its circular genome carrying genes of possible functional importance, are being looked upon as potential drug targets. The genes on this circular genome are believed to be highly conserved among all Plasmodium species. Till date, the plastid genome of P. falciparum, P. berghei and P. chabaudi have been detailed while partial sequences of some genes from other parasites including P. vivax have been studied for identifying evolutionary positions of these parasites. The functional aspects and significance of most of these genes are still hypothetical. In one of our previous reports, we have detailed the complete sequence, as well as structural and functional characteristics of the Elongation factor encoding tufA gene from the plastid genome of P. vivax. We present here the sequences of large and small subunit rRNA (lsu and ssu rRNA) genes, sufB (ORF470) gene, RNA polymerase (rpo B, C) subunit genes and clpC (casienolytic protease) gene from the plastid genome of P. vivax. A comparative analysis of these genes between P. vivax and P. falciparum reveals approximately 5-16% differences. A codon usage analysis of major plastid genes has shown a high frequency of codons rich in A/T at any or all of the three positions in all the species. TTA, AAT, AAA, TAT, and ATA are the major preferred codons. The sequences, functional domains and structural analysis of respective proteins do not show any variations in the active sites. A comparative analysis of these Indian P. vivax plastid genome encoded genes has also been done to understand the evolutionary position of the Indian parasite in comparison to other Plasmodium species., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Thrombocytopenia is commonly seen in Plasmodium vivax malaria, but its prognostic value has not been addressed in children. This prospective study included 676 admitted children of malaria [Plasmodium falciparum (Pf) monoinfection 262, Plasmodium vivax (Pv) monoinfection 380, and mixed (Pf + Pv) infection 34], in which thrombocytopenia (platelet count <150 × 10(3)/mm(3) on admission) was found in 442 (65.38%) children [Pf monoinfection 55.3% (145/262), Pv monoinfection 73.16% (278/380), and mixed infection 55.88% (19/34)]. The association of thrombocytopenia was statistically significant with Pv monoinfection [73.16% (278/380)] in comparison to either Pf monoinfection [55.34% (145/262); odds ratio (OR) = 2.199 (95% confidence interval (CI) 1.577-3.068), p < 0.0001] or mixed infection [55.88% (19/34); OR = 2.152 (95%CI 1.054-4.394), p = 0.032]. In Pv monoinfection, thrombocytopenia was highest in 0-5 years age group and subsequently decreased with advancing age, whereas in Pf monoinfection it was reverse. Severe thrombocytopenia (platelet count <20 × 10(3)/mm(3)) was present in 16.52% (73/442) children [Pv monoinfection 21.58% (60/278) and Pf monoinfection 8.97% (13/145)]. The risk of developing severe thrombocytopenia was also highest in Pv monoinfection [15.79% (60/380)] in comparison to Pf monoinfection [10.59% (13/262); OR = 3.591 (95%CI 1.928-6.690), p < 0.0001]. Bleeding manifestations were associated in 21.27% (94/442) children [Pf monoinfection 9.92% (26/262), Pv monoinfection 16.58% (63/380), and mixed malaria 14.71% (5/34)]. All the children having bleeding manifestations had thrombocytopenia but low platelet counts were not always associated with abnormal bleeding. The association of severe malaria was significantly more among children having Pv monoinfection with platelet counts <20 × 10(3)/mm(3) [OR = 2.569 (95%CI 1.196-5.517), p < 0.014] with specificity of 88.3% and positive predictive value of 85%. Till today, thrombocytopenia is not included in severe malaria criterion described by WHO, but when platelet counts <20 × 103/mm(3), we advocate it to include as one of the severe malaria criteria.

Background & Objectives: Evidence on the community knowledge and perceptions on malaria are crucial to design appropriate health communication strategies for malaria control. Orissa, an Indian state with a large proportion of indigenous populations and hilly terrains contributes to the highest malaria burden in India. A study was undertaken to assess the knowledge on malaria among community members who had experienced fever and chills in the endemic district of Boudh in Orissa., Methods: A cross-sectional community-based survey was carried out with respondents (n=300) who had fever with chills within two weeks prior to data collection through a multi-stage sampling and interviewed them using a pre-tested, structured interview schedule., Results: About 90% of respondents recognized fever as a common symptom of malaria, 72.3% said mosquito bites cause malaria, 70.3% of respondents reported mosquito control and personal protection to be the method of malaria prevention, and 24.6% identified chloroquine as the drug used for treatment. Women and scheduled tribe (ST) respondents were found to have lower level of appropriate knowledge of causes, symptoms, and prevention methods of malaria than their counterparts., Interpretation & Conclusion: The study population had a fair knowledge of malaria about the causes, symptoms, treatment, modes of prevention and outcomes of non-treatment compared to most of the studies conducted in similar settings. However, the relatively low awareness among women and tribal population calls for more context specific communication strategies. Such strategies should be based on information needs assessment of different population subgroups, especially of women and members of the ST community, using media that is accessible and clearly understood by different groups.

The occurrence, relation and magnitude of thrombocytopenia in different species of malaria are not clearly defined. This study included 1,064 patients admitted with malaria to study thrombocytopenia (platelet count <150,000 /cumm) in Plasmodium falciparum (Pf) and Plasmodium vivax (Pv) mono infection and mixed infection (Pf + Pv). The species diagnosis was done by peripheral blood film (PBF) and rapid diagnostic test (RDT). Validation by polymerase chain reaction (PCR) was done only in patients with severe thrombocytopenia (platelet count <20,000 /cumm). The breakup of patients was 525 (49.34%) Pf, 460 (43.23%) Pv and 79 (7.42%) mixed malaria (Pf + Pv). Thrombocytopenia was observed in 24.6% (262/1064) patients. The risk was greatest in the mixed infections in comparison to monoinfection individually (43.04% [34/79]; mixed vs Pv monoinfection: Odds Ratio [OR] = 1.675 [95% Confidence Interval (CI) 1.029-2.726], p < 0.0366; mixed vs Pf monoinfection: OR=3.911 [95% CI 2.367-6.463], p < 0.0001). Pv monoinfection (31.09% [143/460]) had greater risk compared to Pf monoinfection (16.19% [85/525]; OR = 2.335 [95% CI 1.722-3.167], p < 0.0001). The occurrence of severe thrombocytopenia was also higher in Pv monoinfection (18.18% [26/143]) in comparison to either Pf monoinfection (10.59% [9/85], OR = 1.877 (95% CI 0.834-4.223)) or mixed infection (11.76% [4/34]; OR = 1.667 (95% CI 0.540-5.142) but this association was statistically not significant. Six patients (3 Pv, 2 Pf and 1 mixed) developed severe epistaxis requiring platelet transfusion. There was no relation between parasite density and platelet count as many patients with severe thrombocytopenia had parasite density similar to patients without thrombocytopenia. We found that the association of thrombocytopenia was statistically more significant with P. vivax monoinfection as compared to P. falciparum.

Point mutations in the dhfr and dhps genes of Plasmodium falciparum are associated with pyrimethamine and sulfadoxine resistance respectively. In this study we have analyzed these genes from Bikaner (situated in North-West region of India), where both uncomplicated and severe manifestations of P. falciparum malaria are seen. A majority of isolates showed double mutant allele for DHFR. In contrast, the only reported mutation present in DHPS was A437G in few isolates. In addition, three novel non-synonymous mutations were observed in the PfDHPS from this region viz., S587F, N666K and C668W. The mutations at the 666 and 668 codon seem to form a bend in the big loop region of the DHPS enzyme and may affect the binding of the drug to the enzyme. Molecular docking of sulfadoxine to this mutated structure indicates reduction in its binding affinity to this enzyme.

Background & Objectives: To examine the household economic impact of an outbreak of chikungunya in terms of out-of-pocket health care expenditure and income foregone due to loss of productive time in Orissa, India., Methods: Structured interviews were conducted on 150 respondents, bread winners from the affected households of a village with maximum number of reported cases in the state, during August 2007. We looked at the economic profile, treatment history, and patient-side cost of care, loss of productivity and consequent income loss., Results: The median out-of-pocket health care expenditure was US$ 84, of which the proportion of cost of diagnosis was the highest (US$ 77). One hundred and forty nine respondents incurred cost of care more than 10% of their monthly household income (catastrophic health expenditure). The median catastrophic health care expenditure was 37%. The respondents depended more on private health care providers (49%) and 31% of them accessed care from both public and private health care providers. The median work days lost was 35 with a consequent loss of income of US$ 75., Interpretation & Conclusion: Outbreak of an emerging disease creates unforeseen catastrophic health care expenditure and reinforcing the poverty ill-health nexus. The priorities of tackling emerging diseases should include; discretionary public health spending, financial protection against the cost of illness and productivity with special emphasis on people living on daily wages with less financial reserves, and further research on therapeutic measures to reduce the duration of suffering and consequent economic loss.

A number of stage-specific antigens have been characterized for vaccine development against Plasmodium falciparum malaria. This study presents a comprehensive analysis of the sequence polymorphism in Plasmodium falciparum apical membrane antigen-1 (PfAMA-1) in population samples from the eastern and western parts of India. This is the first study of its kind for the nearly full length PfAMA-1 gene from these regions in India. Our observations confirmed that sequence diversity of PfAMA-1 confines only to point mutations and shows 4-8% variation as compared to the prototypes. As opposed to the previous studies on PfAMA-1, our study revealed a greater degree of polymorphism in the Domain II region of PfAMA-1 protein, though signature for diversifying selection is seen throughout the gene. Our present investigation also indicates a very high degree of variation in the reported T- and B-cell epitopes of PfAMA-1. Few noteworthy and unique observations made in this study are the substitution of Cysteine residues responsible for the disulfide bond structure of the protein and the presence of premature termination after 595 amino acids in 3 of the 13 isolates under consideration. These crucial findings add new perspectives to the future of AMA-1 research and could have major implications in establishing AMA-1 as a vaccine candidate.

The Apicomplexan parasite Plasmodium vivax is responsible for causing greater than 50% of human malaria cases in Central and South America, Southeastern Asia and the Indian subcontinent. The rising severity of the disease and the resistance shown by the parasite towards usual therapeutic regimen has put forth a demand for a novel drug target to combat this disease. Apicoplast, an organelle of prokaryotic origin, and its circular genome are being looked upon as a potential drug target. The Apicoplast genome is known to carry various genes of functional importance, including the gene encoding for the protein Elongation factor Tu (tuf A) that participates in the translational process in prokaryotes. The tuf A gene is translationally active within the organelle and is believed to be one of the best functionally conserved protein throughout the species. Till date there are no reports of this gene from another major human malaria parasite P. vivax. This is the first report detailing any complete gene analysis from the Apicoplast genome of the Indian P. vivax isolates. The study predicts and evaluates the complete Apicoplast Elongation factor tuf A gene and EF-Tu protein at primary, secondary and tertiary structure level. In addition, a comparative phylogenetic analysis using this gene is done to understand the evolutionary status of Indian P. vivax isolates. Our study shows that although the Indian P. vivax EF-Tu is not showing any major difference at the structural and predicted functional level, it is diverging way ahead from the P. vivax clade.