High Frequency of Cryptosporidium hominis Infecting Infants Points to A Potential Anthroponotic Transmission in Maputo, Mozambique

Cryptosporidium is one of the most important causes of diarrhea in children less than 2 years of age. In this study, we report the frequency, risk factors and species of Cryptosporidium detected by molecular diagnostic methods in children admitted to two public hospitals in Maputo City, Mozambique. We studied 319 patients under the age of five years who were admitted due to diarrhea between April 2015 and February 2016. Single stool samples were examined for the presence of Cryptosporidium spp. oocysts, microscopically by using a Modified Ziehl-Neelsen (mZN) staining method and by using Polymerase Chain Reaction and Restriction Fragment Length Polymorphism (PCR-RFLP) technique using 18S ribosomal RNA gene as a target. Overall, 57.7% (184/319) were males, the median age (Interquartile range, IQR) was 11.0 (7-15) months. Cryptosporidium spp. oocysts were detected in 11.0% (35/319) by microscopy and in 35.4% (68/192) using PCR-RFLP. The most affected age group were children older than two years, [adjusted odds ratio (aOR): 5.861; 95% confidence interval (CI): 1.532-22.417; p-value < 0.05]. Children with illiterate caregivers had higher risk of infection (aOR: 1.688; 95% CI: 1.001-2.845; p-value < 0.05). An anthroponotic species C. hominis was found in 93.0% (27/29) of samples. Our findings demonstrated that cryptosporidiosis in children with diarrhea might be caused by anthroponomic transmission.

High Frequency of Cryptosporidium hominis Infecting Infants Points to A Potential Anthroponotic Transmission in Maputo, Mozambique.

Cryptosporidium is one of the most important causes of diarrhea in children less than 2 years of age. In this study, we report the frequency, risk factors and species of Cryptosporidium detected by molecular diagnostic methods in children admitted to two public hospitals in Maputo City, Mozambique. We studied 319 patients under the age of five years who were admitted due to diarrhea between April 2015 and February 2016. Single stool samples were examined for the presence of Cryptosporidium spp. oocysts, microscopically by using a Modified Ziehl-Neelsen (mZN) staining method and by using Polymerase Chain Reaction and Restriction Fragment Length Polymorphism (PCR-RFLP) technique using 18S ribosomal RNA gene as a target. Overall, 57.7% (184/319) were males, the median age (Interquartile range, IQR) was 11.0 (7-15) months. Cryptosporidium spp. oocysts were detected in 11.0% (35/319) by microscopy and in 35.4% (68/192) using PCR-RFLP. The most affected age group were children older than two years, [adjusted odds ratio (aOR): 5.861; 95% confidence interval (CI): 1.532-22.417; p-value < 0.05]. Children with illiterate caregivers had higher risk of infection (aOR: 1.688; 95% CI: 1.001-2.845; p-value < 0.05). An anthroponotic species C. hominis was found in 93.0% (27/29) of samples. Our findings demonstrated that cryptosporidiosis in children with diarrhea might be caused by anthroponomic transmission.

Intestinal protozoa in hospitalized under-five children with diarrhoea in Nampula - a cross-sectional analysis in a low-income setting in northern Mozambique.

BACKGROUND: In Mozambique, infection by intestinal parasites is reported all over the country. However, infection in children with diarrhoea is mostly focused in the southern region of Mozambique. This work aims to determine the frequency and potential risk factors for infection by Cryptosporidium spp., Giardia lamblia, and Entamoeba histolytica in children under-five years hospitalized with diarrhoea in Hospital Central de Nampula, northern Mozambique. METHODS: A cross-sectional hospital-based surveillance was conducted between March 2015 and January 2018 in children admitted with diarrhoea in Hospital Central de Nampula. Sociodemographic information was obtained through semi-structured interviews applied to the children's caregivers. A single stool sample was collected from each child to detect antigens from Cryptosporidium spp., G. lamblia, and E. histolytica using an immune-enzymatic technique. Crude and adjusted odds ratios (with 95% Confidence Intervals) were obtained by logistic regression models to identify factors associated with infection by Cryptosporidium spp. and G. lamblia. RESULTS: The median age and interquartile intervals of our sample population was 12 months (8-20). Intestinal protozoa were detected in 21.4% (59/276). Cryptosporidium spp. was the most common protozoa (13.9% - 38/274), followed by G. lamblia (9.1% - 25/274) and E. histolytica (0.4% - 1/275). Children with illiterate caregiver's (p-value = 0.042) and undernourished (p-value = 0.011) were more likely to be infected by Cryptosporidium spp. G. lamblia was more common in children living in households with more than four members (p-value = 0.039). E. histolytica was detected in an eleven month's child, co-infected with Cryptosporidium spp. and undernourished. CONCLUSION: Cryptosporidium spp. and Giardia lamblia were the most common pathogenic intestinal protozoa detected in children with diarrhoea hospitalized in the Hospital Central de Nampula. Our findings obtained highlight the importance of exploring the caregiver's education level, children's nutritional status for infections with Cryptosporidium spp., and living conditions, namely crowded households for infections with G. lamblia in children younger than five years.

Rotavirus A infection in children under five years old with a double health problem: undernutrition and diarrhoea - a cross-sectional study in four provinces of Mozambique.

BACKGROUND: Mozambique has a high burden of group A rotavirus (RVA) infection and chronic undernutrition. This study aimed to determine the frequency and potential risk factors for RVA infection in undernourished children under 5 years old with diarrhoea in Mozambique. METHODS: The analysis was conducted using data from March 2015 to December 2017, regarding children under 5 years old with at least one type of undernutrition. Anthropometric measures were used to calculate indices of weight-for-age, weight-for-height and height-for-age through the Z-Scores. RVA results were extracted from the National Diarrhoea Surveillance database. Descriptive statistics, chi-square test was used for qualitative variables and organized in contingency tables and 95% Confidence Intervals (CI) were considered for the calculation of RVA infection proportion and in the multiple logistic regression models to estimate the adjusted odds ratios (AOR). RESULTS: Of the 842 undernourished children included in the analysis, 27.2% (95% CI: 24.3-30.3%) were positive for RVA. The rate of RVA infection was 42.7% (95% CI: 38.0-47.5%) in the pre-vaccine period, with great reduction to 12.2% (95% CI: 9.4-15.6%) in the post-vaccine period. Most of the RVA undernourished children had severe wasting (33.3%) and severe stunting (32.0%). The risk of infection was significantly high in children from 0 to 11 months (p-value < 0.001) when compared to the age group of 24-59 months. A higher proportion of RVA infection was detected in households with five or more members (p-value = 0.029). Similar proportions of RVA were observed in children fed only by breast milk (34.9%) and breast milk with formula (35.6%). A higher proportion of undernourished HIV-positive children co-infected with RVA (7.4%) was observed. CONCLUSIONS: The frequency of RVA infection in undernourished children declined following the introduction of the vaccine in Mozambique. Beyond the temporal variation, Maputo province, age and crowded households were also associated to RVA infection. A high proportion of RVA infection was observed in children with severe wasting and a triple burden of disease: undernutrition, RVA and HIV, highlighting the need to conduct follow-up studies to understand the long-term impact of these conditions on children's development.

Epidemiologia e caracterização molecular de rotavirus A em crianças menores de cinco anos com diarreia aguda, antes e depois da introdução da vacina em moçambique (2013-2018) / Epidemiology and molecular characterization of rotavirus A in children under five with acute diarrhea, before and after the introduction of the vaccine in Mozambique (2013-2018)

A doença diarreica aguda permanece como um dos principais problemas de saúde pública no mundo, onde cerca de 37% das crianças menores de cinco anos morrem por ano, principalmente, em países com baixa renda. Dentre os diferentes agentes etiológicos, os rotavírus são considerados os principais causadores dessa doença. Duas vacinas já foram licenciadas contra a infecção por rotavírus, a vacina pentavalente RotaTeq e a monovalente Rotarix. Ambas as vacinas têm a capacidade para reduzir a gravidade da doença diarreica em crianças menores de cinco anos. O estudo multicêntrico Entérico Global mostrou que o rotavírus é um dos principais agentes causadores da doença diarreica em Moçambique. A fim de se reduzir a gravidade da infecção poro rotavírus A Moçambique introduziu, em setembro de 2015, no seu Programa Alargado de Vacinação, a vacina monovalente Rotarix. O objetivo deste trabalho foi determinar o impacto da vacina na prevenção da infecção e na diversidade dos genótipos de RVA antes e após a introdução da vacina na cidade de Maputo. Para efeito, foi realizado um estudo descritivo transversal em crianças menores de cinco anos de idade, internadas com diarreia aguda nos Serviços de Pediatria do Hospital Central de Maputo, entre os anos de 2013 e 2018. No estudo, as características epidemiológicas e clínicas das crianças selecionadas foram obtidas através de um formulário padronizado

A identificação dos casos positivos para rotavírus em amostras de fezes foi realizado usando o teste de ELISA. As amostras positivas foram submetidas a genotipagem por semi-nested RT-PCR e, parte das amostras, foram posteriormente submetidas a análise de sequenciamento nucletídico. Os dados recolhidos foram introduzidos na base de dados do pacote estatístico Excel 2010 e analisados através do programa estatístico SPSS versão 20 e pacote estatístico R. Foram viii coletadas 802 amostras de fezes durante o período pré-vacinal (2013 - 2015) e 196 no período pós-vacinal (2016 - 2018), onde observou-se uma redução da mediana de casos de hospitalização por diarreia de 76%, diminuindo de 28,7% (82/284) no período pré-vacinal para 15,5% (10/66) no período pós-vacinal, (p <0,05) em todas as faixas etárias. De forma similar, houve uma redução da mediana de 53,89% na proporção de internação por diarreia por rotavírus em crianças <12 meses de idade, de (28,3%) no período pré-vacinal para 13,04% no período pós-vacinal. Após a introdução da vacina verificou-se alteração dos picos sazonais habitualmente observados. As combinações de genótipos de rotavirus mais prevalentes no período pré-vacinal foram: G2P[4] em 2013,62/86 (72,0%), G1P[8]31/48 (65,0%) em 2014, G9P[8],48 (69,0%) em 2015 e no período pós - vacinal foram: G1P [8] 5/10 (50,0%) em 2016 e 5/8(62,2%) 2017;3G3P[8], G3P[4], G4P[6] 1/3 (33,0%) em 2018Em Moçambique, após a introdução da vacina contra o rotavírus, foi observada uma redução nas hospitalizações associadas a rotavírus em todos os grupos etários com mais destaque em crianças <12 meses de idade. Uma grande diversidade de cepas de rotavírus circulou nos períodos pré e pós-introdução da vacina. Os resultados deste estudo sustentam evidências do impacto da vacinação contra o rotavírus destacando a importância na continuidade do programa assim como nos estudos de monitoramento de genótipos de rotavírus depois da introdução da vacina em Moçambique. (AU)

Prevalência e caracterização molecular de rotavírus em crianças menores de 5 anos com diarreia aguda no distrito de chókwè-moçambique / Prevalence and molecular characterization of rotavirus in children under 5 years with acute diarrhea in chókwè-mozambique district

A diarréia é uma importante causa de morbidade e mortalidade em todas as regiões do mundo e entre todas as idades, ainda pouco se sabe sobre a fracção de episódios de diarréia e óbitos de cada patógeno. Os rotavírus (RVs) foram designados como principal causa de doença diarréica e desihidratação grave em lactentes e crianças menores de 5 anos de idade. O rotavirus é oblíquo. O pico de incidência da doença ocorre no inverno nos países do climas temperado e durante todo nos países tropicais. Infecção RVA responsavel por causar 527.000 mortes por ano em todo o mundo. Na África RVs é responsável por 20% a 40% de todas as doenças diarreicas em crianças com menos de 60 meses de idade. Além disso, na África Subsaariana a mortalidade associada a RVA foi estimada em 145,000 mortes / ano. Em Moçambique não há dados que reportam os genótipos de rotavírus que ocorrem e sua contribuição na etiologia diarréia. O desenvolvimento de vacinas RVs tornou-se uma prioridade de saúde pública nos últimos anos, duas vacinas contra o rotavírus: Rotarix, (GSK, Rixensart, Bélgica) e pentavalente (Merck, NJ, EUA). Rotarix foi demonstrado ser eficaz na prevenção da doença grave causada por estirpes G1P[8], G3P[8], G4P[8], G9P[8] enquanto pentavalente na prevenção de doença causada por estirpes G1, G3, G4 e G9. Neste estudo propusemos a caracterizar as estirpes de RVA que ocorrem em população selecionada de crianças menores de cinco anos com diarréia aguda residente no distrito de Chókwè, Moçambique. A presença de rotavírus foi rastreada por EIA em 254 amostras, e 24% foram positivas rotavírus. A dsRNA foi extraído da suspensão fecal pelo metodo de sílica. A detecção de RVA foi realizada pela técnica de EPGA resultados obtidos estre os dois métodos demonstrou concordância em 75%. Todas as amostras apresentaram perfil eletroforético longo e compactáveis com rotavírus do grupo A, porém nenhuma infecção mista foi observada. A síntese de cDNA a partir do RNA extraído foi realizada utilizando-se o iniciador randômico contido no Kit High Capacity® (Applied Biosystems, USA), A PCR foi realizada com um pares de iniciadores consensuais para VP4 (4con2/4con3) e VP7 (9con1/9con2) onde os produtos da obtidos foram submetidos a PCR semi-nested multiplex usando iniciadores específicos para os genotipos G e P. As amostras que possiveies ser caracterizadas pelo PCR semi-nested foram submetidos a analise de sequencia. As combinações de genótipos predominantes foram RVA G12P [8] (51%); G1P [8] (19%); G12P [6] (6%). As informações produzidas neste estudo iram subsidiar a importância do monitoramento dos genótipos de rotavírus antes e durante a introdução da vacina contra o rotavírus e reforcara achados dos trabalhos dos países vizinhos da emergencia G12 na África para o qual ainda não existe vacina mostra proteção.

Diarrhea is an important cause of morbidity and mortality in all regions of the world and among all ages, yet little is known about the fraction of diarrhea episodes and deaths due to each pathogen. Rotaviruses (RVs) were designated as leading cause of severe diarrheal disease and dehydration in infants and young children (under 5 years of age) throughout the world and it is oblique. Peak incidence of disease is winter seasonality in temperate climates and year-round exposure in tropical countries. RVs infection causes 527,000 deaths per year worldwide. In Africa only RVs is responsible for 20% to 40% of all diarrheal diseases in children under 60 months of age. Furthermore in sub-Saharan Africa the mortality associated with RVs has been estimated to 145.000 deaths/ year. In Mozambique there is no data available reporting genotypes of rotavirus occurring and its contribution in diarrhea etiology. To overcome the burden caused by RVs disease, the development of RVs vaccines became a public health priority In the past years, two rotavirus vaccines: Rotarix, (GSK, Rixensart, Belgium) and RotaTeq (Merck, NJ, USA). Rotarix has been shown to be effective in the prevention of severe RVs disease caused by G1P8, G3P8, G4P8, G9P8 strains while RotaTeq in prevention of RV disease caused by G1, G3, G4 and G9 strains. We proposed to characterize of the RVs strains occurring in in selected population of children under five years with acute diarrhea resident in Chokwe district, Mozambique. The presence of rotavirus was screened by EIA in 254 sample and 24% were Rotavirus positive. dsRNS was extracted from fecal suspension by silica procedures and the RVs detection was also performed by PAGE results from these two diagnostic methods demonstrated agreement in 75%. All samples were showed electrophoretic profile long and compactable with group A rotavirus. No mixed infection was observed. Complementary DNA was obtained by reverse transcription with random primer. PCR were done with a pair of consensus primers for VP4 (4con2/4con3) or VP7 (9con1/9con2) and the products were submitted to Multiplex semi-nested PCR with specific primers for the G and P. The product of PCR was submitted to sequencing. The predominant combinations RVA genotypes were G12P[8] (51%,); G1P[8] (19%);G12P[6] (6%). The information produced in this study will subside the importance of monitoring rotavirus genotypes before and during introducing rotavirus vaccine and it emphasize the emergence of G12 type in Africa for which there is no vaccine shows protection ye