Molecular diagnosis of the curly lettuce parasitic contamination from hydroponic cultivation from supermarkets / Diagnóstico molecular da contaminação parasitária de alface crespa de cultivo hidropônico em supermercados

Abstract The consumption of vegetables has increased in recent years due to the search for a healthier diet that is rich in fiber and has fewer calories. To assess the parasitic contamination of lettuce sold in markets, a survey of parasites was carried out from a supermarket chain in the city of Londrina, Paraná. A total of thirty samples of lettuce were purchased in the ten markets visited, three in each, of which ten were conventionally cultivated, ten were hydroponically cultivated, and ten were organically cultivated. All samples were analyzed using the sedimentation methods of Hoffman, Pons and Janer and the fluctuation method of Faust and colleagues and Willis with adaptations. In addition, the samples were subjected to DNA extraction by a commercial kit and polymerase chain reaction to detect Toxoplasma gondii, Cryptosporidium spp. and Giardia spp., which are protozoa that cause food and waterborne parasitic outbreaks. All samples were negative for sedimentation and flotation techniques. One of the hydroponically cultivated samples was positive for T. gondii. The results demonstrate the risk of curly lettuce contamination from hydroponic cultivation and the need for proper cleaning of these foods before consumption.

Resumo O consumo de vegetais aumentou nos últimos anos devido à busca de uma dieta mais saudável, rica em fibras e com menos calorias. Para avaliar a contaminação parasitária de alface vendida nos mercados, foi realizado um levantamento de parasitas em vegetais folhosos de uma cadeia de supermercados da cidade de Londrina, Paraná. Um total de 30 amostras foram compradas nos dez mercados visitados, três em cada, dos quais dez foram convencionalmente cultivados, dez cultivados hidroponicamente e dez foram cultivados organicamente. Todas as amostras foram analisadas, usando-se os métodos de sedimentação de Hoffman, Pons e Janer e o método de flutuação de Faust e colaboradores e Willis com adaptações. Além disso, as amostras foram submetidas à extração de DNA por um kit comercial e reação em cadeia da polimerase (PCR) para detectar Toxoplasma gondii, Cryptosporidium spp. e Giardia spp., que são protozoários causadores de surtos transmitidos pela água e alimentos. Todas as amostras foram negativas para técnicas de sedimentação e flutuação. Uma das amostras cultivadas hidroponicamente foi positiva para T. gondii. Os resultados demonstram o risco de contaminação por alface crespa do cultivo hidropônico e a necessidade de limpeza adequada desses alimentos antes do consumo.

Secondary transmission of cryptosporidiosis associated with well water consumption: two case studies

Abstract: Cryptosporidiosis is a very prominent disease in the field of public health, and usually causes diarrhea. We describe two immunocompetent patients who presented with chronic diarrhea that was ultimately found to be caused by continuous exposure to well water contaminated with the microbial cysts (oocysts) of the Cryptosporidium spp parasite. We describe the patients' histories and possible explanations for their prolonged symptoms.

Infección por Cryptosporidium parvum en una mujer embarazada, inmunocompetente, con riesgo ocupacional / Cryptosporidium parvum infection in a pregnant inmunocompetent woman with occupational risk

La criptosporid iosis es una zoonosis parasitaria provocada por diversas especies de Cryptosporidium. Esta coccidiosis afecta a múltiples vertebrados, incluido el ser humano. En Chile, al igual que en otros países, es una infección poco frecuente en inmunocompetentes y adquiere gran relevancia en pacientes inmunocom-prometidos. Se presenta el caso de una egresada de la carrera de Medicina Veterinaria, embarazada, con 20 semanas de gestación, procedente del sector de Laguna Verde, Región de Valparaíso, que fuera infectada por Cryptosporidium sp. El diagnostico etiológico se realizó con tinción de Ziehl Neelsen, RPC anidada y posterior secuenciación. En el mismo periodo se detectó la infección en sus gatos asintomáticos. En ella y los animales se identificó a C. parvum. Su cónyuge así como sus otras mascotas no estaban infectados. Este corresponde al primer reporte de una posible transmisión de criptos-poridiosis entre ser humano y gato.

Cryptosporidioses is a parasitic zoonoses generated by diverse Cryptosporidium species. This coccidiosis affects multiple vertebrate species, including human beings. In Chile, as it happens in other countries, cryptosporidioses is a low frequency infection in immunocompetent individuals, acquiring a big relevance in immunocompromised ones. We present the following case: a recently graduated student from Veterinary medical school, with a 20 week pregnancy, living in “Laguna Verde” area in the Region of Valparaíso and who was infected with Cryptosporidium sp. Etiologic diagnosis was made by Ziehl Neelsen, and nested PCR followed by PCR product sequencing. During the same period, the infection was detected in her cats which were asymptomatic. In all of them, her and the cats, the species identified was Cryptosporidium parvum. Her husband and her other pets were all asymptomatic and non infected. This is the first report of a possible cryptosporidioses transmission between humans and cat.

[Cryptosporidium genetic diversity based on analysis of COWP, SSU-rRNA and TRAP-C2 polymorphic genes in children with diarrhea in Tehran and qazvin provinces: a survey from Iran]

Since Cryptosporidium is a worldwide distributed protozoan parasite and is considered as one of the most common causes of infection and diarrhea in humans with autoimmune deficiency, as well as in young live stock, molecular epidemiologic studies of cryptosporidiasis will be helpful for underlying transmission and molecular pathogenesis of Cryptosporidium in humans. The aim of the present study was to determine the species and genotypes of Cryptosporidium among children with diarrhea in Tehran and Qazvin provinces by PCRRFLP using the three polymorphic regions of SSU-rRNA, COWP and TRAP-C2 genes. 1263 stool samples were collected from the children less than 12 years with diarrhea who referred to Pediatrics Medical Centers in Gazvin and Tehran Provinces, Iran, during 2005-2007. After determination of the presence of Cryptosporidium oocytes by ZiehlNeelsen acid, fast staining genomic DNA was extracted. Nested PCR-RFLP was performed by -rRNA, COWP and TRAP-C2 genes. Results of microscopically positive samples showed that the overall prevalence of infection in children was 31 [2.5%]. Results of nested PCR amplification showed that of 31 isolates of children, all of three targeted gene were successfully amplified. Our results indicated that the zoonotic transmission is the main mode of infection in Iran and indicates that direct or indirect contact with animals, especially calf, is possibly the main route of human infection

Molecular characterization of Cryptosporidium spp. from HIV infected patients from an urban area of Brazil / Caracterização molecular de Cryptosporidium spp. de pacientes de área urbana do Brasil infectados por HIV

Cryptosporidium spp. are important cause of enteric disease in humans, but may also infect animals. This study describes the relative frequency of several Cryptosporidium species found in human specimens from HIV infected patients in the São Paulo municipality obtained from January to July 2007. Sequence analysis of the products of nested-PCR based on small subunit rRNA and Cryptosporidium oocyst wall protein coding genes revealed 17 (63.0 percent) isolates of C. hominis, four (14.8 percent) C. parvum, five (18.5 percent) C. felis and one (3.7 percent) C. canis. These findings suggest that, in urban environments of Brazil, the cat adapted C. felis may play a potential role in the zoonotic transmission of cryptosporidiosis whereas the anthroponotic transmission of cryptosporidiosis caused by C. hominis seems to predominate.

Cryptosporidium spp. são importantes causas de doenças entéricas em humanos, mas podem também ser encontrados em animais. O presente estudo descreve a frequência relativa de diversas espécies de Cryptosporidium em amostras de humanos da cidade de São Paulo, Brasil, obtidas de janeiro a julho de 2007. Análises de sequências de produtos de nested PCR direcionadas ao genes codificadores da menor unidade ribosomal e da proteina de parede de oocistos revelaram 17 (63,0 por cento) isolados de C. hominis, quatro (14,8 por cento) C. parvum, cinco (18,5 por cento) C. felis, e um (3,7 por cento) C. canis. Estes resultados sugerem que, em ambientes urbanos no Brasil, o genótipo adaptado ao gato pode desempenhar potencial papel na transmissão zoonótica de criptosporidiose, enquanto a transmissão antroponótica da criptosporidiose causada pelo C. hominis parece predominar.

Environmental risk factors for Cryptosporidium infection in an island from Western Venezuela

Few investigations have been conducted on risk factors for Cryptosporidium infection in communities from developing countries. A study was conducted to determine the prevalence and risk factors for cryptosporidiosis in San Carlos island, Venezuela. A sample of 515 subjects (mean age ± SD: 21.4 ± 17.8 years) was surveyed. Single fecal specimens were collected and modified Ziehl-Neelsen carbolfuchsin staining of formalin-ether concentrate stools were examined for identification of the parasite. Infections with Cryptosporidium (67 of 515, 13 percent) were common. Prevalence of the parasite varied among sectors of the community; 34 of 67(50.7 percent) cases of cryptosporidiosis clustered in two sectors with extreme poverty. Variables strongly associated with a higher risk for the infection (p < 0.01) were residing in these sectors versus the remainder, living in a hut or small residence versus a brick or larger house, using an area of backyard rather than a toilet or latrine for defecation, and having contact with soil contaminated with human feces. Crowding was also a risk (p < 0.05). Contact with human feces contaminated-soil may be an important mode of transmission and poverty a predisposing factor for the infection.

Genetic characterization of cryptosporidium spp. among children with diarrhea in Tehran and Qazvin Provinces, Iran

Cryptosporidium is an intracellular apicomplexan parasite that infects a wide range of vertebrates including humans. Cryptosporidiosis is a major cause of diarrhea in children with and without human immunodeficiency virus [HIV] infection in developing countries. More recently, the molecular methods for identification of morphologically indistinguishable species have been developed. The aim of this study was to determine the characterization of various species of this coccidian among children with diarrhea by using molecular methods. Fecal samples were collected from 1263 children with diarrhea who referred to Pediatrics Medical Centers in Qazvin and Tehran, two central provinces of Iran. Initial identification of Cryptosporidium was carried out by Zeihl-Neel-sen acid-fast staining method of stool samples. DNA was extracted from positive microscopically samples and were subjected to a two step nested PCR-RFLP based on SSU-rRNA gene. Out of 1263 collected samples, 31 [2.5%] were found to be contained Cryptosporidium oocysts. RFLP analysis showed that 80.6% of the positive isolates were Cryptosporidium parvum, 16.1% C. hominis and 3.2% had mix infection pattern of both C. parvum and C hominis. Our results showed that the zoonotic pattern of transmission is predominant and has considerable significance in epidemiology of Cryptosporidiosis in the study areas

Cryptosporidium spp. em alimentos / Cryptosporidium spp. in food

O Cryptosporidium parvum foi descrito pela primeira vez em 1907 por Tyzzer, porém reconhecido como patógeno humano apenas em 1976. A Cryptosporidiose, causada principalmente pelo Cryptosporidium parvum, é mais grave em crianças, idosos e indivíduos imunodeficientes ou imunossuprimidos. Em contrapartida, em imunocompetentes, a doença é auto-limitante, caracterizada por uma diarréia aquosa que pode durar de 3 a 14 dias sem maiores complicações. O Cryptosporidium parvum é transmitido por oocistos, por meio de contato direto com as fezes de pessoas e animais infectados ou indiretamente durante a ingestão de água e alimentos contaminados. Os oocistos do Cryptosporidium spp. São resistentes aos desinfetantes químicos usados no tratamento da água de bebida, são facilmente destruídos pela fervura, mas não são retidos pelos filtros domésticos convencionais; porém, os filtros com poros com menos de 1 micrômetro de diâmetro e de osmose reversa são eficazes na retenção dos oocistos presentes na água. Os esgotos lançados (in natura) poluem os recursos hídricos, principalmente na falta de sistemas de captação e tratamento adequados. Estudos recentes demonstram que nos USA, 65-97 por cento das águas de superfície (rios, lagos, etc) estão contaminados com oocistos de Cryptosporidium spp, o que representa um sério risco ao aparecimento de surto de cryptosporidiose por meio da contaminação do sistema de abastecimento de água potável e da contaminação de frutas e verduras que são consumidas sem receber tratamento adequado. O surto de cryptosporidiose humana com maior repercussão, ocorreu na cidade de Milwaukee - Wisconsin - USA, envolvendo cerca de 300 mil pessoas, com um custo total de 90,2 milhões de dolares gastos no tratamento e controle da doença. O presente trabalho de revisão tem como objetivo principal alertar as autoridades e órgãos competentes para um maior controle microbiológico da água de abastecimento público e dos alimentos.

nested-PCR assay for detection of cryptosporidium parvum oocysts in water samples

Cryptosporidiosis is a gastroenteric disease caused by the protozoan parasite Cryptosporidium parvum. Water-borne transmission of this organism has become more prevalent in recent years. Current method for detection of C. parvum oocysts in water is immunofluoresence assay [IFA]. The method is time consuming, laborious and particularly not-specific. It cannot determine the infectivity of detected oocysts. We have evaluated a nested- PCR assay for sensitive detection of C. parvum oocysts in water samples. Water sample concentrates were spiked with Cryptosporidium oocysts and after DNA extraction and purification by QIAamp DNA mini kit, detection was achieved by nested PCR amplification of a 200 bp region of hsp70 gene specific for C. Parvum. The method could detect as few as one oocyst in seeded tap water samples. On the basis of these results, PCR could be a useful tool in the monitoring of water samples for the detection of Cryptosporidium oocysts

Acción de distintos coagulantes para la eliminación de Cryptosporidium spp. en el proceso de potabilización del agua / The action of different coagulants to remove Cryptosporidium during the process of water treatment

Cryptosporidium es uno de los microorganismos de mayor interés desde el punto de vista de la Salud Pública y constituye un problema prioritario para las plantas y organismos reguladores de agua. Debido a su pequeño tamaño y a su resistencia a la cloración, la eliminación por el proceso de potabilización es una tarea compleja. En este trabajo se analizó la efectividad de distintos coagulantes utilizados comúnmente en tal proceso para lograr la remoción de los ooquistes. Se trabajó con la prueba de jarras (Jar Test). Se halló que: 1) Los coagulantes con agregado de polímeros coadyuvantes producen remociones de ooquistes superiores a 2 log. 2) Un valor bajo de turbiedad no asegura una remoción óptima de los parásitos. 3) La adición de polielectrolitos al cloruro férrico disminuye la variabilidad tanto en la turbiedad final como en la eliminación de Cryptosporidium.

Cryptosporidium is one of the microorganisms of main concern from the point of view of Public Health, being a priority problem for water treatment plants and water regulatory institutions. Due to its small size and resistance to chlorination, Cryptosporidium removal during the process of drinking water treatmentis a hard task. The effectiveness of different coagulants commonly used in the process of removal of oocysts was analyzed. Thetechnique used was the Jar Test. It was found that: 1) coagulants with the addition of polimeric coadjuvants produce over 2 logs of oocyst removal; 2) a low value in turbidity does not necessarily mean optimal parasite removal, and 3) the addition of polyelectrolites to ferric chloride diminishes variability, both in final turbidity and Cryptosporidium removal.

El desafío de las enfermedades emergentes y reemergentes / The challenge of emerging and re-emerging diseases

En los albores del siglo XXI, las enfermedades emergentes o reemergentes plantean un serio desafío para su control. Las enfermedades emergentes son enfermedades infecciosas con incidencia en aumento en las últimas dos décadas. Se identifican con diversas etiologías: priónica, viral, bacteriana, micótica o parasitaria; en diferentes poblaciones de riesgo, con diferentes vías de transmisión, historia natural y eco epidemiología. Como reemergentes figuran aquellas afecciones infecciosas conocidas que reaparecen después de una disminución significativa de su incidencia. Para controlar estos preocupantes fenómenos, se debe interpretar a estas dolencias infecciosas dentro de una ecología tanto global como local con carácter dinámico, pues están moduladas por cambios tecnológicos, sociales, económicos, ambientales y demográficos, a lo que se suma el potencial de cambio y capacidad de adaptación de los microorganismos. El desafío requiere una estrategia integrada, que contemple el fortalecimiento de la vigilancia epidemiológica y del laboratorio, con alto nivel científico, así como la construcción de infraestructura internacional y local capaz de responder con soluciones adecuadas y oportunas. Es imprescindible el desarrollo de estrategias de prevención y control de las enfermedades emergentes, por medio de intervenciones eficaces basadas en investigaciones prácticas y realistas. El objetivo de esta revisión no exhaustiva sobre el tema es encarar las generalidades de estas infecciones, profundizando en algunos ejemplos seleccionados

Updates on AIDS crytosporidiosis: a review

Cryptosporidio en un parásito que se caracteriza entre otras enfermedades por la formación de diarreas crónicas acuosos. El curso de la enfermedad depende mayormente del estado inmunológico del paciente. Las diarreas pueden causar hasta un 80% de mortalidad en pacientes inmunosuprimidos como de SIDA. Aunque hasta el momento no existe ningún tipo de terapia efectiva, resultados prometedores se le han atribuido al uso de Espiromicina, Eritromicina, Somatostatina y sus análogos, y a Zidovudina

Cryptosporidiosis intestinal: un caso de infección intrahospitalaria / Intestinal cryptosporidiosis: a case of nosocomial infection

En un lactante con diarrea prolongada, en el cual la investigación para cryptosporidium habría sido previamente negativa, se detectó el parásito en coincidencia con la admisión, a la misma sala del hospital, de otro niño que sufría diarrea aguda y en cuyas deposiciones se identificaron ooquistes de cryptosporidium sp. La evidencia sugiere transmisión intrahospitalaria del agente