We present filaria-nested polymerase chain reaction (PCR)

Previously, Plasmodium knowlesi was not considered as a species of Plasmodium that could cause malaria in human beings, as it is parasite of long-tailed (Macaca fascicularis) and pig-tailed (Macaca nemestrina) macaques found in Southeast Asia. A case of infection by P. knowlesi is described in a Spanish traveller, who came back to Spain with daily fever after his last overseas travel, which was a six-month holiday in forested areas of Southeast Asia between 2008 and 2009. His P. knowlesi infection was detected by multiplex Real time quantitative PCR and confirmed by sequencing the amplified fragment. Using nested multiplex malaria PCR (reference method in Spain) and a rapid diagnostic test, the P. knowlesi infection was negative. This patient was discharged and asymptomatic when the positive result to P. knowlesi was reported. Prior to this case, there have been two more reports of European travellers with malaria caused by P. knowlesi, a Finnish man who travelled to Peninsular Malaysia during four weeks in March 2007, and a Swedish man who did a short visit to Malaysian Borneo in October 2006. Taken together with this report of P. knowlesi infection in a Spanish traveller returning from Southeast Asia, this is the third case of P. knowlesi infection in Europe, indicating that this simian parasite can infect visitors to endemic areas in Southeast Asia. This last European case is quite surprising, given that it is an untreated-symptomatic P. knowlesi in human, in contrast to what is currently known about P. knowlesi infection. Most previous reports of human P. knowlesi malaria infections were in adults, often with symptoms and relatively high parasite densities, up to the recent report in Ninh Thuan province, located in the southern part of central Vietnam, inhabited mainly by the Ra-glai ethnic minority, in which all P. knowlesi infections were asymptomatic, co-infected with P. malariae, with low parasite densities and two of the three identified cases were very young children under five years old.

Background Strongyloides stercoralis is an intestinal parasite that can cause chronic infection, hyperinfection and/or a dissemination syndrome in humans. The use of techniques targeting ova fails to detect S. stercoralis, as only larvae of the parasite are excreted in faeces. Due to the absence of “Gold” standard diagnostic method for S. stercoralis, there is a paucity of reported data worldwide. Objective This study aimed to evaluate the performance of diagnostic methods of S. stercoralis infection by taking the composite reference as a “Gold” standard. Methods A cross-sectional study was conducted among 844 schoolchildren in Amhara Region, Ethiopia, from April to December 2019. Stool samples were collected and processed with formol-ether concentration technique (FECT), spontaneous tube sedimentation technique (STST), Baermann concentration technique (BCT), agar plate culture (APC) and real-time polymerase chain reaction (RT-PCR). Sensitivity, specificity, positive predictive value, and negative predictive value of each diagnostic method were computed against the composite reference. The agreements of diagnostic methods were evaluated by Kappa value at 95% CI. Results The composite detection rate of S. stercoralis by the five diagnostic methods was 39.0% (329/844). The detection rate of the parasite from stool samples by FECT, STST, BCT, APC and RT-PCR was 2.0% (17/844), 4.0% (34/844), 10.2% (86/844), 10.9% (92/844) and 28.8% (243/844), respectively. The highest detection rate (37.8%; 319/844) of S. stercoralis was recorded by a combination of BCT, APC, and RT-PCR followed by a combination of STST, BCT, APC and RT-PCR (37.3%; 315/844). The sensitivity of FECT, STST, BCT, APC and RT-PCR against the composite reference was 5.2%, 10.3%, 26.4%, 28.0% and 73.9%, respectively. The diagnostic agreements of RT-PCR, APC, BCT, STST and FECT with the composite reference in detection of S. stercoralis were substantial (0.775), fair (0.321), fair (0.305), slight (0.123), and slight (0.062), respectively. Conclusion RT-PCR detected the highest number of S. stercoralis infections. A combination of RT-PCR with APC and/or BCT better detected S. stercoralis from stool samples compared to other combinations or single diagnostic methods. Therefore, RT-PCR and combination of RT-PCR with APC and/or BCT diagnostic methods should be advocated for detection of S. stercoralis infection.

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Background: Malaria is one of the deadliest disease in the world, particularly in Africa. The resistance to antimalarial drugs is one of the most important problems to global malaria control. The study assess the evolution of the different resistance markers over time and the possible influence of the interventions and treatment changes that have been made in Equatorial Guinea.Methods: 1223 biological samples distributed from 1999 to 2019 were included in the study. The screening of the mutations in pfdhfr, pfdhps, pfmdr1 and pfcrt genes were carried out by nested PCR and RFLPs, and the study of pfk13 genes was carried out by nested PCR, followed by sequencing to determine the presence of mutations Results: the haplotypes partially and fully resistant (pfdhfr + pfdhps) increase over time. In 2019, the fully resistant is increasing although super resistant remains lower. There is also a continued decline over time in pfmdr1 and pfcrt gene mutations. Since 2008, when ACTs were introduced in the country, the number of mutations detected in pfk13 has been increasing over time, with more mutations being observed in 2019. In this year, were detected 2 synonymous and 5 non-synonymous mutations, although are not related to resistance to ACTs. In addition, the non-synonymous A578S mutation, the most frequent on the African continent, was detected in 2013, although it has not been detected in the following years.Conclusions: The withdrawal of the use of CQ as a treatment in the country has been effective over time, as wild-type parasite populations outnumber mutant populations. The upward trend observed in SP resistance markers evidence its misuse alone or in combination with AS or AQ in some areas of the country, this allows selective pressure from SP to continue. SNPs 540E and 581G do not exceed the limit of 50% and 10% respectively, which means that SP as an IPT is still effective in the country. As for the pfk13 gene, no mutations have been detected in relation to resistance to ACTs. However, in 2019 there is a greater accumulation of non-synonymous mutations compared to years prior to 2008.

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