Characterisation of Anopheles species composition and genetic diversity in Meghalaya, northeast India, using molecular identification tools.

Malaria in India is declining, in part due to the use of long-lasting insecticide-treated nets (LLINs) and vector control. Historically, the north-eastern region of India has contributed ~10%-12% of the nation's malaria burden. The important mosquito vectors in northeast India have long been considered to be Anopheles baimaii and An. minimus, both associated with forest habitats. Local deforestation and increased rice cultivation, along with widespread LLIN use, may be changing vector species composition. Understanding if and how vector species composition is changing is critical to successful malaria control. In Meghalaya state, malaria is now at a low level of endemicity with occasional seasonal outbreaks. In a biodiverse setting like Meghalaya, where >24 Anopheles mosquito species have been recorded, accurate morphological identification of all species is logistically challenging. To accurately determine Anopheles species richness in the West Khasi Hills (WKH) and West Jaintia Hills (WJH) districts, adult and larval mosquitoes were collected and identified using molecular methods of allele-specific PCR and cytochrome oxidase I DNA barcoding. In 14 villages across both districts, we identified high species richness, 19 species in total. Molecular findings indicated that An. minimus and An. baimaii were rare, while four other species (An. maculatus, An. pseudowillmori, An. jeyporiensis and An. nitidus) were abundant. Anopheles maculatus was highly prevalent in WKH (39% of light trap collections) and An. pseudowillmori in WJH (45%). Larvae of these four species were found in rice fields, suggesting that land cover change is influencing species composition change. Our results suggest that rice fields might be contributing to the observed abundance of An. maculatus and An. pseudowillmori, which could be playing a role in malaria transmission, either independently due to their high abundance, or in combination with An. baimaii and/or An. minimus.

Antimalarial Drug Resistance Profiling of Plasmodium falciparum Infections in India Using Next-Generation Sequencing.

Background: Tracking the emergence and spread of antimalarial drug resistance has become critical to sustaining progress towards the control and eventual elimination of malaria in South Asia, especially India. Methods: An amplicon sequencing protocol was used for high-throughput molecular surveillance of antimalarial drug resistance in a total of 158 isolates at three sites in India: Chennai, Nadiad and Rourkela. Five genes of the Plasmodium falciparum implicated in antimalarial resistance were investigated here; Pfcrt for chloroquine resistance, Pfdhfr for pyrimethamine resistance, Pfdhps for sulfadoxine resistance, Pfk13 for artemisinin resistance and Pfmdr1 for resistance to multiple antimalarials. Results: Mutations in the propeller domain of PfK13 were observed in two samples only, however these mutations are not validated for artemisinin resistance. A high proportion of parasites from the P. falciparum dominant site Rourkela showed wild-type Pfcrt and Pfdhfr haplotypes, while mutant Pfcrt and Pfdhfr haplotypes were fixed at the P. vivax dominant sites Chennai and Nadiad. The wild-type PfDHPS haplotype was predominant across all study sites. Finally, we observed the largest proportion of suspected multi-clonal infections at Rourkela, which has the highest transmission of P. falciparum among our study sites. Conclusion: This is the first simultaneous high-throughput next generation sequencing of five complete P. falciparum genes from infected patients in India.

Comparison of the effectiveness of Gingival unit transfer and free Gingival graft in the management of localized Gingival recession - A systematic review.

Background: Gingival Recession (GR) is defined as the displacement of the soft tissue margin apical to the cementoenamel junction which can lead to root exposure and hypersensitivity. Treatment of GR has become an important therapeutic issue due to the increasing number of cosmetic requests from patients. Several techniques exist for the management of GR that include Sub-Epithelial Connective Tissue Graft (SECTG), Pedicle Graft (lateral and coronal), and Free Gingival Graft (FGG) and more. FGG is a non-submerged grafting procedure carried out for the management of recession defects. However, FGG has limitations like aesthetic mismatch and bulky appearance. A relatively newer modification of FGG was introduced by Allen in 2004 wherein a palatal graft including the marginal gingiva and interdental tissue was used as donor tissue for recession coverage. This review aims to study and compare the use of Gingival Unit Graft/Transfer (GUG/GUT) (palatal graft including the marginal gingiva and papillae) and FGG in the management of GR. Materials and Methods: Randomized Clinical Trials, Non-Randomized Controlled Clinical Trials for the treatment of Miller Class I,II, and III of GRs by GUG with FGG were identified. Data sources included electronic databases and hand-searched journals. The primary outcome variables were complete root coverage, mean root coverage, vertical recession depth. The secondary outcome variables were keratinized tissue width gain, clinical attachment level and probing depth. Results: Three Randomized Control Trials (RCTs) met the inclusion criteria and were evaluated in this systematic review. Both techniques showed significant improvement in clinical parameters. GUG procedure resulted in a greater percentage of sites achieving complete root coverage and vertical recession depth reduction when compared to FGG group in all the studies. Two studies reported significantly greater mean root coverage in GUG group compared to FGG group. GUG procedure revealed statistically significant greater gain in keratinized tissue width when compared to FGG group in all the studies. Conclusion: Because of the limited number of selected studies, no conclusive statement could be made regarding the advantage of the GUG technique over FGG. However, the percentage of sites with complete root coverage obtained in the GUG technique is higher than FGG. More RCTs with aesthetic and patient satisfaction-related parameters are needed to provide definite evidence.

Systematic review of school-based tobacco prevention programs for the adolescents in India from 2000 to 2020.

In India, 14.6% of adolescents are currently using tobacco in any form and tobacco control is a major public health challenge. The objective of this systematic review is to analyze all the existing literature and evaluate the effectiveness of school-based tobacco use prevention programs for adolescents in India. The review protocol was registered in PROSPERO (CRD42020159535). Studies were selected using database search, manual search, gray literature, reference chasing, and contacting the authors. All randomized controlled trials, cluster-randomized trials, quasi-experimental, and non-randomized studies reporting school-based tobacco use preventive interventions for adolescents in India; articles published in English (other languages where it can be translated to English) published between January 2000 till May 2020 were included. Data was independently extracted by two reviewers. The Risk of bias (RoB) and quality of the study were assessed using appropriate tools. Among 7972 identified articles, only 13 studies met the inclusion criteria. Each study implemented a unique intervention and measured distinct outcomes. Postintervention, all the studies reported improvements in the study group with respect to the reduction of tobacco use and change in the knowledge, attitude, practices, and/or behavior outcome parameters. Twelve study results were based on short-term assessment. Overall, a 5.17-17.0% tobacco use reduction rate was noted. RoB was high for six studies. Key methodological problems related to study design, duration, outcome parameter, follow-up time, type of intervention, and attrition were identified. School-based tobacco use prevention programs for adolescents in India might have shown positive outcomes but are associated with significant limitations.

Population genetic structure of the malaria vector Anopheles fluviatilis species T (Diptera: Culicidae) in India.

Anopheles fluviatilis James (Diptera: Culicidae) represents a complex that comprises four sibling species (S, T, U, and V). Among these, species T is widely distributed in India. Chromosomal inversion polymorphism exists among different geographic populations of An. fluviatilis species T; however, population genetic structure is not understood. This study inferred a genetic structure among six geographically diverse populations of species T using a panel of microsatellite markers. Analyses indicated a significant but low genetic differentiation among the majority of the studied populations. A significant correlation was observed between genetic and geographic distances, exhibiting stepwise migration patterns among populations.

Genetic diversity in two leading Plasmodium vivax malaria vaccine candidates AMA1 and MSP119 at three sites in India.

Plasmodium vivax, a major contributor to the malaria burden in India, has the broadest geographic distribution and shows higher genetic diversity than P. falciparum. Here, we investigated the genetic diversity of two leading P. vivax vaccine candidate antigens, at three geographically diverse malaria-endemic regions in India. Pvama1 and Pvmsp119 partial coding sequences were generated from one hundred P. vivax isolates in India (Chennai n = 28, Nadiad n = 50 and Rourkela n = 22) and ~1100 published sequences from Asia, South America, North America, and Oceania regions included. These data were used to assess the genetic diversity and potential for vaccine candidacy of both antigens on a global scale. A total of 44 single nucleotide polymorphism (SNPs) were identified among 100 Indian Pvama1 sequences, including 10 synonymous and 34 nonsynonymous mutations. Nucleotide diversity was higher in Rourkela and Nadiad as compared to Chennai. Nucleotide diversity measures showed a strong balancing selection in Indian and global population for domain I of Pvama1, which suggests that it is a dominant target of the protective immune response. In contrast, the Pvmsp119 region showed highly conserved sequences in India and across the Oceania, South America, North America and Asia, demonstrating low genetic diversity in the global population when compared to Pvama1. Results suggest the possibility of including Pvmsp119 in a multivalent vaccine formulation against P. vivax infections. However, the high genetic diversity seen in Pvama1 would be more challenging for vaccine development.

Obstructive sleep apnea domains: Knowledge, attitude and practice results of dentists from a dental college in India.

OBJECTIVE: To assess the knowledge, attitude and practice regarding different domains of obstructive sleep apnea (OSA) amongst dentists from a dental college in India. METHODS: 112 dentists participated in the study. A 23-item, self-designed, pre-tested and validated questionnaire assessing the knowledge, attitude and practice (KAP) regarding nine different domains was used to collect the data. Knowledge and practice was categorized domain wise as good/poor, while attitude as favorable/unfavorable. RESULTS: Dentists were observed to have good knowledge about OSA, for domains concerning to the definition (60.71%), general findings (76.19%) and risk factors of OSA (66.96%). For the domain of screening and diagnosis (38.83%) along with treatment and referral (36.01%) dentists presented poor knowledge. 100% favorable attitude was reported for all the domains, while the dentists poorly faired (<50%) for both the practice domains. CONCLUSION: In spite of dentists showcasing favorable attitude towards OSA, they possessed poor knowledge for domains concerning screening, diagnosis and treatment modalities of OSA which may be linked to the hurdle in their way of practice. Thus a special attention towards these domains needs to be given so as to improve the handling skills of dentist for OSA patients coming to their clinics and prevent further health related issues.

Prevalence of dental caries among children aged 5-15 years from 9 countries in the Eastern Mediterranean Region: a meta-analysis.

BACKGROUND: Dental caries affects all age groups, although children are affected to a greater extent. Several studies have assessed the prevalence of dental caries in the World Health Organization (WHO) Eastern Mediterranean Region. However, prevalence data for dental caries have not been pooled for all countries in the Region. AIMS: To estimate by meta-analysis the pooled prevalence of dental caries among children aged 5-15 years in the Region. METHODS: The study protocol was registered in PROSPERO with registration number CRD42016037157. Twenty-one studies (37 estimates) were identified through systematic search for articles published between 1 January 2005 and 5 July 2018. The required data from each article were extracted into the datasheet. A random-effects meta-analysis was performed for the overall age group and for age 5, 12 and 15 years individually. RESULTS: Data were available for only 9 of the 21 countries in the Region. Heterogeneity between studies was high (I²>98%). There was considerable variation among the countries for the prevalence of dental caries at different ages. The pooled prevalence for deciduous dentition in children aged 5 years was 65% (45-85%); 61% (50-72%) for permanent dentition in children aged 12 years; 70% (64-75%) for children aged 15 years; and 66% (59-73%) for children aged 6-15 years. The most common index used for oral examination to calculate caries experience was WHO Basic Oral Health Survey Criteria, 1997. CONCLUSIONS: Dental caries continues to be an oral health concern among children in 9 countries in the Region.

Effectiveness of licorice in preventing dental caries in children: A systematic review.

BACKGROUND: Dental caries is one of the most prevailing oral health diseases in children. Recent times have focused on herbal products, because they have minimum or no side effects and are effective in prevention. Licorice is one such product belonging to Glycyrrhiza family used in the form of dentifrice, chewing gums, lollipop, gels, etc., Literature reports about the activity of licorice root extract on the biofilm thereby reducing Streptococcus mutans (SM) count and preventing dental caries in children. OBJECTIVES: The objective is to assess the effectiveness of licorice in reducing SM count and preventing dental caries in children. METHODOLOGY: PubMed and Google scholar were searched with search strategies for studies reporting licorice as intervention in children among 3-15 years for preventing dental caries. Only those studies with study design of randomized controlled trials, clinical trials, and comparative studies published between January 1, 2000, and October 31, 2019 were included. Cross references and hand searching for the relevant articles were also conducted. RESULTS: A preliminary search yielded a total of 31 studies through PubMed and Google scholar. From 31 studies, nine studies were excluded based on the screening through titles. From the remaining 22 articles, six duplicates, four without parameters, six as reviews and case reports and were excluded. Finally, six articles giving ten estimates were included for qualitative synthesis. CONCLUSION: Licorice extracts proves to be effective as an antimicrobial agent by reducing the count of SM in children. Its action on biofilm limits the fall of pH thereby preventing acidic environment that increases the risk of dental caries. Moreover, licorice in lollipop form is well accepted by children.

Antibody responses within two leading Plasmodium vivax vaccine candidate antigens in three geographically diverse malaria-endemic regions of India.

BACKGROUND: Identifying highly immunogenic blood stage antigens which can work as target for naturally acquired antibodies in different eco-epidemiological settings is an important step for designing malaria vaccine. Blood stage proteins of Plasmodium vivax, apical membrane antigen-1 (PvAMA-1) and 19 kDa fragment of merozoite surface protein (PvMSP-119) are such promising vaccine candidate antigens. This study determined the naturally-acquired antibody response to PvAMA-1 and PvMSP-119 antigens in individuals living in three geographically diverse malaria endemic regions of India. METHODS: A total of 234 blood samples were collected from individuals living in three different eco-epidemiological settings, Chennai, Nadiad, and Rourkela of India. Indirect ELISA was performed to measure human IgG antibodies against recombinant PvAMA-1 and PvMSP-119 antigens. The difference in seroprevalence and factors associated with antibody responses at each site was statistically analysed. RESULTS: The overall seroprevalence was 40.6% for PvAMA-1 and 62.4% for PvMSP-119. Seroprevalence to PvAMA-1 was higher in Chennai (47%) followed by Nadiad (46.7%) and Rourkela (27.6%). For PvMSP-119, seroprevalence was higher in Chennai (80.3%) as compared to Nadiad (53.3%) and Rourkela (57.9%). Seroprevalence for both the antigens were found to be higher in Chennai where P. vivax is the dominant malaria species. In addition, heterogeneous antibody response was observed for PvAMA-1 and PvMSP-119 antigens at each of the study sites. Two factors, age and malaria positivity were significantly associated with seropositivity for both the antigens PvAMA-1 and PvMSP-119. CONCLUSION: These data suggest that natural acquired antibody response is higher for PvMSP-119 antigen as compared to PvAMA-1 antigen in individuals living in three geographically diverse malaria endemic regions in India. PvMSP-119 appears to be highly immunogenic in Indian population and has great potential as a malaria vaccine candidate. The differences in immune response against vaccine candidate antigens in different endemic settings should be taken into account for development of asexual stage based P. vivax malaria vaccine, which in turn can enhance malaria control efforts.

The burden of submicroscopic and asymptomatic malaria in India revealed from epidemiology studies at three varied transmission sites in India.

Malaria in India, while decreasing, remains a serious public health problem, and the contribution of submicroscopic and asymptomatic infections to its persistence is poorly understood. We conducted community surveys and clinic studies at three sites in India differing in their eco-epidemiologies: Chennai (Tamil Nadu), Nadiad (Gujarat), and Rourkela (Odisha), during 2012-2015. A total of 6,645 subject blood samples were collected for Plasmodium diagnosis by microscopy and PCR, and an extensive clinical questionnaire completed. Malaria prevalence ranged from 3-8% by PCR in community surveys (24 infections in Chennai, 56 in Nadiad, 101 in Rourkela), with Plasmodium vivax dominating in Chennai (70.8%) and Nadiad (67.9%), and Plasmodium falciparum in Rourkela (77.3%). A proportional high burden of asymptomatic and submicroscopic infections was detected in community surveys in Chennai (71% and 71%, respectively, 17 infections for both) and Rourkela (64% and 31%, 65 and 31 infections, respectively). In clinic studies, a proportional high burden of infections was identified as submicroscopic in Rourkela (45%, 42 infections) and Chennai (19%, 42 infections). In the community surveys, anemia and fever were significantly more common among microscopic than submicroscopic infections. Exploratory spatial analysis identified a number of potential malaria hotspots at all three sites. There is a considerable burden of submicroscopic and asymptomatic malaria in malarious regions in India, which may act as a reservoir with implications for malaria elimination strategies.

Assessment of mother's ability in caries diagnosis, utilizing the smartphone photographic method.

BACKGROUND: Smartphone is proved to be reliable in teledentistry model. The aim of the study was to assess the mother's ability in caries diagnosis, utilizing the smartphone-based photographic method. HYPOTHESIS: Mothers have the ability for caries diagnosis utilizing the smartphone photographic method. METHODS: A cohort of 100 mothers was given health education regarding dental caries diagnosis. Then, their children from the preschools were screened by the dentist for dental caries using visual examination, followed by clicking intraoral photographs using Motorola 3G phone. The photographs were sent through WhatsApp to respective mothers, and they were asked to diagnose their child's dental status. Accuracy and reliability measures of mother's diagnosis were estimated by calculating the sensitivity, specificity, predictive values, and kappa value. RESULTS: The calculated value for sensitivity was 88.3%, specificity was 98.3%, positive predictive value was 92%, negative predictive value was 97%, and accuracy was 96%. There was a strong agreement between mother's and dentist's diagnosis (kappa value of 0.87). CONCLUSION: Following the dental health education, it can be concluded that mothers are in a better position to diagnose their child's dental status through smartphone-based photographs.

Dental caries prevalence among 5- to 15-year-old children from SEAR countries of WHO: A systematic review and meta-analysis.

OBJECTIVES: The aim of this review was to estimate the prevalence of dental caries in children 5-15 years of age in the countries of the South-East Asia Region (SEAR) of World Health Organization (WHO) and to describe the different caries indices used in these population-based studies. MATERIALS AND METHODS: A systematic search was carried out in two databases from 1st January 2005 to 31st May 2015. Studies were included if they met the predetermined eligibility criteria. Quality assessment was done with eight-item checklist. Meta-analysis was done for 5, 12, 15, and 6-15 years age group using software STATA version 12. RESULTS: The search strategy yielded 265 unique articles of which 36 met the inclusion criteria included for the review. Data were available for only three SEAR countries. The quality of the majority of the studies ranged from moderate to high. Heterogeneity between the studies was high (I2 > 98%). Variation in dental caries prevalence was found among different ages and among different SEAR countries. The most commonly used index for measuring dental caries was the dentition status of the 1997 WHO criteria. CONCLUSION: Dental caries continues to be a prominent oral health problem among children in the SEAR countries with huge variation in the prevalence across ages and countries. This review results can be used to update the "WHO Oral Health Country/Area Profile Program" for dental caries among children for SEAR.

Assessment of oral findings of dental patients who screen high and no risk for obstructive sleep apnea (OSA) reporting to a dental college - A cross sectional study.

To assess the oral findings of patients who screen high and no risk for obstructive sleep apnea (OSA) reporting to outpatient department of a dental college. Methods: Patients coming to dental Out Patient Department (OPD) were screened using STOP questionnaire and were categorized into high (n=130) and no risk (n=130) OSA groups. BANG (body mass index, age, neck circumference and gender) was recorded for both the OSA risk group patients. Following this oral and general examination was performed using predetermined criteria for temporomandibular disorder (TMD), Angle's Class of Malocclusion, maxillary arch constriction, facial profile, Mallampati score for uvula, tongue size, depth of palatal vault and periodontitis. Chi-squared statistics was applied to know the significant difference among the two groups. Multivariate logistic regression model was run by including the significant variables. Results: 94 females and 166 males were present in the study with a mean age of 43.67±11.89 in both the risk groups. All the variables except Angle's class of malocclusion and periodontitis showed significant difference among high and no risk OSA groups. Logistic regression confirmed that neck circumference, Class 3 or 4 Mallampati score, large tongue and deep palatal vault were commonly observed among high risk OSA group and were independent risk factors for developing high risk of OSA. Conclusion: Neck circumference>40cm, large tongue, Class 3 or 4 Mallampati score and deep palatal vault were found to be independent predictors of developing high risk of OSA. Dentist can play a vital role in screening such patients as he comes in close vicinity of oral cavity and thus can refer the patients to sleep physician to promote interdisciplinary approach.

A Method for Amplicon Deep Sequencing of Drug Resistance Genes in Plasmodium falciparum Clinical Isolates from India.

A major challenge to global malaria control and elimination is early detection and containment of emerging drug resistance. Next-generation sequencing (NGS) methods provide the resolution, scalability, and sensitivity required for high-throughput surveillance of molecular markers of drug resistance. We have developed an amplicon sequencing method on the Ion Torrent PGM platform for targeted resequencing of a panel of six Plasmodium falciparum genes implicated in resistance to first-line antimalarial therapy, including artemisinin combination therapy, chloroquine, and sulfadoxine-pyrimethamine. The protocol was optimized using 12 geographically diverse P. falciparum reference strains and successfully applied to multiplexed sequencing of 16 clinical isolates from India. The sequencing results from the reference strains showed 100% concordance with previously reported drug resistance-associated mutations. Single-nucleotide polymorphisms (SNPs) in clinical isolates revealed a number of known resistance-associated mutations and other nonsynonymous mutations that have not been implicated in drug resistance. SNP positions containing multiple allelic variants were used to identify three clinical samples containing mixed genotypes indicative of multiclonal infections. The amplicon sequencing protocol has been designed for the benchtop Ion Torrent PGM platform and can be operated with minimal bioinformatics infrastructure, making it ideal for use in countries that are endemic for the disease to facilitate routine large-scale surveillance of the emergence of drug resistance and to ensure continued success of the malaria treatment policy.