Plasmodium infection and its association with biochemical and haematological parameters in free-living Alouatta guariba clamitans (Cabrera, 1940) (Primates: Atelidae) in Southern Brazil

BACKGROUND The influence of Plasmodium spp. infection in the health of Southern brown howler monkey, Alouatta guariba clamitans, the main reservoir of malaria in the Atlantic Forest, is still unknown. OBJECTIVES The aim of this study was to investigate the positivity rate of Plasmodium infection in free-living howler monkeys in an Atlantic Forest fragment in Joinville/SC and to associate the infection with clinical, morphometrical, haematological and biochemical alterations. METHODS Molecular diagnosis of Plasmodium infection in the captured monkeys was performed by Nested-polymerase chain reaction (PCR) (18S rRNA and coxI). Haematological and biochemical parameters were compared among infected and uninfected monkeys; clinical and morphometrical parameters were also compared. FINDINGS The positivity rate of Plasmodium infection was 70% among forty captured animals, the highest reported for neotropical primates. None statistical differences were detected in the clinical parameters, and morphometric measures comparing infected and uninfected groups. The main significant alteration was the higher alanine aminotransferase (ALT) levels in infected compared to uninfected monkeys. MAIN CONCLUSIONS Therefore, Plasmodium infection in howler monkeys may causes haematological/biochemical alterations which might suggest hepatic impairment. Moreover, infection must be monitored for the eco-epidemiological surveillance of malaria in the Atlantic Forest and during primate conservation program that involves the animal movement, such as translocations.

Serological and molecular techniques applied for identification of Plasmodium spp. in blood samples from nonhuman primates / Técnicas sorológicas e moleculares aplicadas na identificação de Plasmodium spp. em amostras de primatas não humanos

Abstract The aim of this study was to identify Plasmodium spp. in blood samples from nonhuman primates (NHPs) in the state of Maranhão, using classical and alternative techniques for examination of human malaria. A total of 161 blood samples from NHPs were analyzed: 141 from captive animals at a Wildlife Screening Center (CETAS) and 20 from free-living animals in a private reserve. The techniques used were microscopy, rapid diagnostic test (RDT), Indirect fluorescent antibody test (IFAT) and molecular techniques (semi-nested PCR, quantitative real-time PCR and LAMP). Two serological methods (dot-ELISA and indirect ELISA) were also standardized with rhoptry protein-soluble antigen of P. falciparum and P. berghei. Trophozoite forms of Plasmodium sp. were identified on slides from five different animals. No samples were positive through RDT and LAMP. Four samples were seropositive for P. malariae through IFAT. The samples showed low reactivity to ELISA. Plasmodium sp. was detected in 34.16% (55/161) of the samples using qPCR based on the 18S rRNA gene. After sequencing, two samples showed 100% identityl to P. malariae, one showed 97% identity to Plasmodium sp. ZOOBH and one showed 99% identity to P. falciparum . PCR was shown to be the most sensitive technique for diagnosing Plasmodium in NHP samples.

Resumo Neste estudo objetivamos identificar Plasmodium spp. em amostras sangue de primatas não humanos (PNH) do estado do Maranhão, utilizando técnicas clássicas e alternativas para o exame da malária humana. Foram analisadas 161 amostras de sangue de PNH, sendo 141 de CETAS (cativeiro) e 20 de reserva particular (vida livre), utilizando microscopia, teste de diagnóstico rápido (RDT), imunofluorescência indireta (IFI) e técnicas moleculares (semi-nested PCR, PCR em tempo real quantitativo e LAMP). Dois métodos sorológicos (dot-ELISA e ELISA indireto) também foram padronizados com antígenos solúveis de roptrias de P. falciparum e P. berghei. Formas trofozoíticas de Plasmodium sp. foram identificadas em lâminas de cinco animais diferentes. Nenhuma amostra foi positiva em TDR e LAMP. Quatro amostras foram soropositivas para P. malariae na IFI. Os soros de PNH mostraram baixa reatividade pelo ELISA indireto. Plasmodium sp. foi detectado em 34,16% (55/161) das amostras utilizando a qPCR baseada no gene 18S rRNA. No sequenciamento, duas amostras mostraram identidade com P. malariae (100%), uma com Plasmodium sp. ZOOBH (97%) e uma com P. falciparum (99%). A PCR mostrou ser a técnica mais sensível para diagnósticos de Plasmodium em amostras de PNH.