SARS-CoV-2 Point Mutation and Deletion Spectra, and Their Association with Different Disease Outcome

Mutant spectra of RNA viruses are important to understand viral pathogenesis, and response to selective pressures. There is a need to characterize the complexity of mutant spectra in coronaviruses sampled from infected patients. In particular, the possible relationship between SARS-CoV-2 mutant spectrum complexity and disease associations has not been established. In the present study, we report an ultra-deep sequencing (UDS) analysis of the mutant spectrum of amplicons from the nsp12 (polymerase)- and spike (S)-coding regions of thirty nasopharyngeal isolates (diagnostic samples) of SARS-CoV-2 of the first COVID-19 pandemic wave (Madrid, Spain, April 2020) classified according to the severity of ensuing COVID-19. Low frequency mutations and deletions, counted relative to the consensus sequence of the corresponding isolate, were overwhelmingly abundant. We show that the average number of different point mutations, mutations per haplotype and several diversity indices was significantly higher in SARS-CoV-2 isolated from patients who developed mild disease than in those associated with moderate or severe disease (exitus). No such bias was observed with RNA deletions. Location of amino acid substitutions in the three dimensional structures of nsp12 (polymerase) and S suggest significant structural or functional effects. Thus, patients who develop mild symptoms may be a richer source of genetic variants of SARS-CoV-2 than patients with moderate or severe COVID-19. IMPORTANCEThe study shows that mutant spectra of SARS-CoV-2 from diagnostic samples differ in point mutation abundance and complexity, and that significantly larger values were observed in virus from patients who developed mild COVID-19 symptoms. Mutant spectrum complexity is not a uniform trait among isolates. The nature and location of low frequency amino acid substitutions present in mutant spectra anticipate great potential for phenotypic diversification of SARS-CoV-2.

Community-based approaches for malaria case management in remote communities in the Brazilian Amazon.

INTRODUCTION: Malaria case management is a pivotal intervention in malaria elimination. However, many remote areas in Brazil still lack access to basic health services. This study describes a community-based approach (CBA) for malaria case management in the large remote area of the Jaú National Park (JNP), Amazonas, Brazil. METHODS: In 2001, a general health CBA was initiated with a motor group (MG); a participative community health diagnosis (PCHD) was subsequently implemented between 2001 and 2005. In 2006, a CBA for malaria case management started with an expanded MG including all sectors with a stake in malaria control, from the local residents to the federal government. In 2008, community microscopists were selected and trained to diagnose hemoparasites. A full malaria strategy was implemented in 2009 with subsequent quality control follow-up. RESULTS: Two educational materials were co-created with local communities. The MG identified malaria as a major health problem and the malaria MG planned the control activities. Ten communities selected a resident to become malaria microscopists, and ten solar-operated health centers were built. The number of slide readings increased from 923 in 2006 to 1,900 in 2009, while malaria infections decreased from 354 cases in 2005 to 20 cases in 2015. The excess time (≥ 48 hours) between first symptoms and diagnosis/treatment decreased from 68.9% of cases in 2005 to 14.3% in 2010. CONCLUSIONS: While many factors were likely involved in the reduction of malaria transmission in the JNP, the CBA played an important role in the sustained success of the initiative.

Community-based approaches for malaria case management in remote communities in the Brazilian Amazon

Abstract INTRODUCTION Malaria case management is a pivotal intervention in malaria elimination. However, many remote areas in Brazil still lack access to basic health services. This study describes a community-based approach (CBA) for malaria case management in the large remote area of the Jaú National Park (JNP), Amazonas, Brazil. METHODS In 2001, a general health CBA was initiated with a motor group (MG); a participative community health diagnosis (PCHD) was subsequently implemented between 2001 and 2005. In 2006, a CBA for malaria case management started with an expanded MG including all sectors with a stake in malaria control, from the local residents to the federal government. In 2008, community microscopists were selected and trained to diagnose hemoparasites. A full malaria strategy was implemented in 2009 with subsequent quality control follow-up. RESULTS Two educational materials were co-created with local communities. The MG identified malaria as a major health problem and the malaria MG planned the control activities. Ten communities selected a resident to become malaria microscopists, and ten solar-operated health centers were built. The number of slide readings increased from 923 in 2006 to 1,900 in 2009, while malaria infections decreased from 354 cases in 2005 to 20 cases in 2015. The excess time (≥ 48 hours) between first symptoms and diagnosis/treatment decreased from 68.9% of cases in 2005 to 14.3% in 2010. CONCLUSIONS While many factors were likely involved in the reduction of malaria transmission in the JNP, the CBA played an important role in the sustained success of the initiative.

The Malaria System MicroApp: A New, Mobile Device-Based Tool for Malaria Diagnosis.

BACKGROUND: Malaria is a public health problem that affects remote areas worldwide. Climate change has contributed to the problem by allowing for the survival of Anopheles in previously uninhabited areas. As such, several groups have made developing news systems for the automated diagnosis of malaria a priority. OBJECTIVE: The objective of this study was to develop a new, automated, mobile device-based diagnostic system for malaria. The system uses Giemsa-stained peripheral blood samples combined with light microscopy to identify the Plasmodium falciparum species in the ring stage of development. METHODS: The system uses image processing and artificial intelligence techniques as well as a known face detection algorithm to identify Plasmodium parasites. The algorithm is based on integral image and haar-like features concepts, and makes use of weak classifiers with adaptive boosting learning. The search scope of the learning algorithm is reduced in the preprocessing step by removing the background around blood cells. RESULTS: As a proof of concept experiment, the tool was used on 555 malaria-positive and 777 malaria-negative previously-made slides. The accuracy of the system was, on average, 91%, meaning that for every 100 parasite-infected samples, 91 were identified correctly. CONCLUSIONS: Accessibility barriers of low-resource countries can be addressed with low-cost diagnostic tools. Our system, developed for mobile devices (mobile phones and tablets), addresses this by enabling access to health centers in remote communities, and importantly, not depending on extensive malaria expertise or expensive diagnostic detection equipment.

Epidemiology of imported malaria among children and young adults in Barcelona (1990-2008).

BACKGROUND: Increasing international travel and migration is producing changes in trends in infectious diseases, especially in children from many European cities. The objective of this study was to describe the epidemiology and determine the trends of imported malaria in patients under 20 years old in the city of Barcelona, Spain, during an 18-year period. METHODS: The study included malaria cases that were laboratory confirmed and reported to the malaria register at the Public Health Agency of Barcelona from 1990 to 2008, residing in Barcelona and less than 20 years old. Patients were classified as natives (born in Spain) or immigrants. Differences in the distribution of demographic, clinical characteristics, and incidence per 100,000 person-year evolution were analysed. Natives and immigrants were compared by logistic regression by calculating the odds ratio (OR) with a 95% confidence interval (CI) and Chi-square for a linear trend (p<0.05). RESULTS: Of the total 174 cases, 143 (82.1%) were immigrants, 100 (57.5%) were female, 121 (69.5%) Plasmodium falciparum, and 108 (62.1%) were visiting friends and relatives (VFR) as the reason for travel. Among the immigrants, 99 (67.8%) were from Equatorial Guinea. Immigrant cases more frequently travelled to Africa than natives (p=0.02). The factors associated with imported malaria among immigrant residents was travelling for VFR (OR: 6.2 CI 1.9-20.2) and age 15-19 (OR: 3.7 CI 1-13.3). The incidence increased from 1990 to 1999 (p<0.001) and decreased from 2000 to 2008 (p=0.01), although the global linear trend was not statistically significant (p=0.41). The fatality rate was 0.5%. CONCLUSIONS: The majority of cases of malaria in population less than 20 years in Barcelona were immigrants, travelling to Africa for VFR and Plasmodium falciparum was most frequently detected. The trend analysis of the entire study period did not show a statistically significant decline. It is recommended to be aware of malaria, especially among children of immigrants who travel to their parent's home country for VFR. Better access to pre travel advice should be provided.

Imported malaria among African immigrants: is there still a relationship between developed countries and their ex-colonies?

BACKGROUND: The objective of this study was to compare cases of imported malaria originating from the Spanish ex-colony of Equatorial Guinea (EG) with those originating from the rest of Africa (RA). METHODS: All the African cases detected in Barcelona between 1989 and 2007 were investigated in a retrospective analysis. Clinical-epidemiological variables such as sex, age, visiting friends and relatives (VFR), species, hospital admission and chemo-prophylaxis were compared. Data were analysed by logistic regression, calculating the Odds Ratio (OR) and 95% Confidence Intervals (95% CI). RESULTS: Of the 489 African patients, 279 (57,1%) had been born in EG and 210 (42,9%) in the rest of Africa. The cumulative incidence of imported malaria among those from EG was 179.6 per thousand inhabitants, while in those from the RA it was 33.7 per thousand (p < 0.001). Compliance with chemoprophylaxis (CP) was very low, but there were no differences between the two groups. Comparing those from EG to those from RA, the former were characterized by having more patients in the visiting friends and relatives (VFR) category, and more individuals younger than 15 years or older than 37 years, and more women. They also visited a traveller's health centre more often, had fewer hospital admissions and were less likely to reside in the inner city. CONCLUSION: Cases of imported malaria originating in Africa, are more likely to come from the Spanish ex-colony of EG, and VFR are more likely to be affected. It is recommended that developed countries promote prevention programmes, such as CP advice directed at African immigrants, and develop programmes of cooperation against malaria in their ex-colonies.

A tuberculose nas populaçöes indígenas das Américas: com uma visäo especial para as populaçöes indígenas do Norte da Amazônia, uma revisäo da literatura / The tuberculosis in the indigenous populations in the Americas: with a special vision for the indigenous populations in the north of the Amazônia: review literature

Encontramos evidências da existência de tuberculose como doença humana nas Américas já desde a época pré-colombiana. Näo acontece o mesmo na Amazônia. Este fato levou a explicar a gravidade da situaçäo atual das populaçöes indígenas da Amazônia basicamente pela falta de contato prévio com o agente etiológico. O objetivo é de, através de uma revisäo da literatura, discutir o comportamento atual nos grupos indígenas do norte da Amazônia. Foram revisados um total de 21 trabalhos das Américas contendo um total de 130 casos de tuberculose prehistórica. A maior parte dos trabalhos da América do Sul säo estudos de caso, ao contrário da América do Norte em que séries populacionais foram analisadas. As principais localizaçöes dos achados estäo no Chile, no Perú, nas pradarias centrais da América do Norte e próximo ao lago Ontário. O material na maioria dos casos é ósseo, embora na América do Sul encontramos 16 casos em múmias. A forma respiratória da doença foi encontrada em 30 casos, sendo 19 deles inferidos a partir de lesöes costais, dos restantes, 105-110 foram formas ósseas e 26 outras localizaçöes. Como determinantes mais importantes encontrou-se as mudanças em certas práticas culturais e condiçöes de vida, as quais poderiam ter atuado em ciclos de agravamento. Também parece ter existido facilidade para a expansäo da endemia pela mobilidade e intensidade de contatos intergrupais. Da Amazônia foram revisados um total de 14 trabalhos, centrando a discussäo em cinco do Alto Rio Negro e da Area Indígena Yanomami. Os trabalhos foram basicamente estudos clínicos nas aldeias, tendo sido utilizadas diferentes metodologias para efetuar o diagnóstico, diferenciando entre pessoas infectadas e doentes. Näo säo muitos os dados encontrados em relaçäo à idade, sexo e localizaçäo das patologias, embora observa-se como dado interessante a existência de grande número de localizaçöes extrapulmonares. Como determinantes mais importantes encontramos que, em todos os casos, a endemizaçäo da tuberculose parece ser funçäo das mudanças contextuais ocorridas, como mudanças de práticas culturais, de economia, de arquitetura de habitaçöes, maiores agrupamentos e confinamento dos indivíduos, assim como a existência de deslocamento de contingentes humanos, e contatos interétnicos.