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Acanthamoebae spp. are considered the most commonly occurring free-living amoebae (FLA) in the environment. Their high resilience enables them to thrive in different types of environments. Using purposive sampling, 80 surface water samples were collected from identified coastal sites in Mariveles, Bataan, and Lingayen Gulf (40 water samples for each). Nineteen (23.75%) of the 80 water samples yielded positive amoebic growth during the 14-day culture and microscopic examination. The polymerase chain reaction confirmed Acanthamoeba spp. DNA in isolates MB1, A3, A4, A7, C5, and D3 using JDP1 and JDP2 primer sets. Further sequencing revealed that the isolates belonged to Acantha- moeba sp., Acanthamoeba culbertsoni, Acanthamoeba castellani, and Acanthamoeba genotype T4. The sequences were deposited in GenBank and registered under accession numbers PP741651, PP767364, PP741728, PP741729, PP767365, and PP767366, respectively. Potential risk factors such as waste disposal, expansion of human settlements to coastal locations, and soil runoffs in these environments should be controlled to mitigate the proliferation of potentially pathogenic strains of FLAs. [ABSTRACT FROM AUTHOR]

Natural hot springs are ideal places and environmental matrices that offer relaxation to people and microorganisms of different types. A total of 40 surface water samples were collected from the five identified collection sites, eight water samples for each site. Collection sites are designated 200 m apart to cover the entire study site. Surface water samples were collected approximately 10–20 cm from the surface. Water samples were filtered, cultured, and microscopically observed for 14 days. After 14 days of cultivation, eight (20%) water samples revealed cystic and trophozoite stages. Polymerase chain reaction using JDP1 and JDP2 specific primers confirmed the presence of Acanthamoeba spp. from two of our isolates in the hot spring, isolates 1.1 and 5.1. Further sequencing revealed that the isolates are Acanthamoeba T20 and Acanthamoeba genotype T7. Sequences were deposited to GenBank and were assigned accession numbers PP741726 and PP741727, respectively. The isolation of Acanthamoeba spp. in hot springs has significant health implications, especially for those who use it for recreational activity. Private resort owners are highly encouraged to regularly monitor and maintain hot spring resorts to avoid future infections. [ABSTRACT FROM AUTHOR]

Research on free-living amoebae (FLA) and its public health implication as an etiologic agent of parasitic infection in humans has recently gained traction in the Philippines. This study aimed to identify potential FLAs in collected groundwater samples from Masinloc, Zambales, Philippines. Fifty-four (54) water samples were collected in 250-mL sterile polyethylene containers by purposive sampling from selected groundwater sources in six (6) barangays of Masinloc. The samples were vacuum filtered through a 1.2-µm pore glass microfiber filter, cultured onto non-nutrient agar (NNA) lawned with Escherichia coli, and observed microscopically for amoebic growth for 14 days using light microscopy. Amoebic growth was observed in 11.1% (6/54) of water samples. DNAs from positive samples were extracted and were made to react with polymerase chain reaction using Acanthamoeba-specific JDP1 (5'-GGCCCAGATCGTTTACCGTGAA-3') and JDP2 (5'-TCTCA-CAAGCTGCTAGGGAGTCA-3') primers, and universal primer Euk A (5'-AACCTGGTTGATCCTGCCAGT-3') and Euk B (5'-TGATCCTTCTGCA GGTTCACCTAC-3'). The presence of Acanthamoeba genotypes T4, T7, and T11 was confirmed using molecular and phylogenetic analysis. Our results confirmed that groundwater sources from two of six sampling sites (33.3%) in Masinloc, Zambales, were contaminated with potentially pathogenic FLAs. Proper identification of risk factors that may cause contamination consequently leads to the implementation of programs that will prevent future infections. [ABSTRACT FROM AUTHOR]

The dual effects of co-infection of Plasmodium spp. and hookworm on malaria remain under debate. This study investigated prevalence, prevalence odds ratio (POR) of co-infection and impact of co-infection on malaria parasite density and haemoglobin levels in comparison to Plasmodium mono-infection. The protocol for this systematic review and meta-analysis is registered at PROPERO under ID: CRD42020202156. Relevant literatures were obtained from PubMed, ISI Web of Science, and Scopus on 25 December 2020. Mean difference (MD) and confidence interval (CI) of malaria parasite density and haemoglobin were compared using a random effect model. Heterogeneity was assessed using Cochrane Q and I2 statistics. Publication bias was determined by visualising funnel plot asymmetry. Of 1756 articles examined, 22,191 malaria cases across 37 studies included 6096 cases of co-infection of Plasmodium spp. and hookworm. The pooled prevalence was 20% (95% CI 15–26%, I2 99.6%, 37 studies) and was varied in terms of geographical region. Co-infection occurred by chance (OR 0.97, p 0.97, 95% CI 0.73–1.27, I2 95%, 30 studies). The mean malaria parasite density for co-infection (478 cases) was similar to Plasmodium mono-infection (920 cases) (p 0.24, MD 0.86, 95% CI − 0.58–2.29, I2 100%, 7 studies). The mean haemoglobin level for co-infection (90 cases) was similar to Plasmodium mono-infection (415 cases) (p 0.15, MD − 0.63, 95% CI − 1.49–0.23, I2 98%, 4 studies). Co-infection was common and occurred by chance but varied by geographic region. Further studies are required to investigate the mechanism of hookworm infection on malaria severity. Additionally, detection of hookworm infections among patients with malaria in endemic areas of both diseases is recommended to prevent severe malaria. [ABSTRACT FROM AUTHOR]

A better understanding of the occurrence and risk of Plasmodium vivax infection among Duffy-negative individuals is required to guide further research on these infections across Africa. To address this, we used a meta-analysis approach to investigate the prevalence of P. vivax infection among Duffy-negative individuals and assessed the risk of infection in these individuals when compared with Duffy-positive individuals. This study was registered with The International Prospective Register of Systematic Reviews website (ID: CRD42021240202) and followed Preferred Reporting Items for Systematic review and Meta-Analyses guidelines. Literature searches were conducted using medical subject headings to retrieve relevant studies in Medline, Web of Science, and Scopus, from February 22, 2021 to January 31, 2022. Selected studies were methodologically evaluated using the Joanna Briggs Institute (JBI) Critical Appraisal Tools to assess the quality of cross-sectional, case–control, and cohort studies. The pooled prevalence of P. vivax infection among Duffy-negative individuals and the odds ratio (OR) of infection among these individuals when compared with Duffy-positive individuals was estimated using a random-effects model. Results from individual studies were represented in forest plots. Heterogeneity among studies was assessed using Cochrane Q and I2 statistics. We also performed subgroup analysis of patient demographics and other relevant variables. Publication bias among studies was assessed using funnel plot asymmetry and the Egger's test. Of 1593 retrieved articles, 27 met eligibility criteria and were included for analysis. Of these, 24 (88.9%) reported P. vivax infection among Duffy-negative individuals in Africa, including Cameroon, Ethiopia, Sudan, Botswana, Nigeria, Madagascar, Angola, Benin, Kenya, Mali, Mauritania, Democratic Republic of the Congo, and Senegal; while three reported occurrences in South America (Brazil) and Asia (Iran). Among studies, 11 reported that all P. vivax infection cases occurred in Duffy-negative individuals (100%). Also, a meta-analysis on 14 studies showed that the pooled prevalence of P. vivax infection among Duffy-negative individuals was 25% (95% confidence interval (CI) − 3%–53%, I2 = 99.96%). A meta-analysis of 11 studies demonstrated a decreased odds of P. vivax infection among Duffy-negative individuals (p = 0.009, pooled OR 0.46, 95% CI 0.26–0.82, I2 = 80.8%). We confirmed that P. vivax infected Duffy-negative individuals over a wide prevalence range from 0 to 100% depending on geographical area. Future investigations on P. vivax infection in these individuals must determine if Duffy-negativity remains a protective factor for P. vivax infection. [ABSTRACT FROM AUTHOR]

Human African trypanosomiasis (HAT) is endemic in Africa; hence, the possibility of co-infection with malaria among patients with HAT exists. The present study investigated co-infection with malaria among patients with HAT to provide current evidence and characteristics to support further studies. Potentially relevant studies that reported Plasmodium spp. infection in patients with HAT was searched in PubMed, Web of Science, and Scopus. The risk of bias among the included studies was assessed using the checklist for analytical cross-sectional studies developed by the Joanna Briggs Institute. The pooled prevalence of Plasmodium spp. infection in patients with HAT was quantitatively synthesized using a random-effects model. Subgroup analyses of study sites and stages of HAT were performed to identify heterogeneity regarding prevalence among the included studies. The heterogeneity of the outcome among the included studies was assessed using Cochran's Q and I2 statistics for consistency. Publication bias was assessed if the number of included studies was 10 or more. For qualitative synthesis, a narrative synthesis of the impact of Plasmodium spp. infection on the clinical and outcome characteristics of HAT was performed when the included studies provided qualitative data. Among 327 studies identified from three databases, nine studies were included in the systematic review and meta-analysis. The prevalence of Plasmodium spp. co-infection (692 cases) among patients with HAT (1523 cases) was 50% (95% confidence interval [CI] = 28–72%, I2 = 98.1%, seven studies). Subgroup analysis by type of HAT (gambiense or rhodesiense HAT) revealed that among patients with gambiense HAT, the pooled prevalence of Plasmodium spp. infection was 46% (95% CI = 14–78%, I2 = 96.62%, four studies), whereas that among patients with rhodesiense HAT was 44% (95% CI = 40–49%, I2 = 98.3%, three studies). Qualitative syntheses demonstrated that Plasmodium spp. infection in individuals with HAT might influence the risk of encephalopathy syndrome, drug toxicity, and significantly longer corrected QT time. Moreover, longer hospital stays and higher treatment costs were recorded among co-infected individuals. Because of the high prevalence of malaria among patients with HAT, some patients were positive for malaria parasites despite being asymptomatic. Therefore, it is suggested to test every patient with HAT for malaria before HAT treatment. If malaria is present, then antimalarial treatment is recommended before HAT treatment. Antimalarial treatment in patients with HAT might decrease the probability of poor clinical outcomes and case fatality in HAT. [ABSTRACT FROM AUTHOR]

Co-infection with malaria and human immunodeficiency virus (HIV) increases the severity and mortality rates of both diseases. A better understanding of the effects of co-infections could help in the diagnosis, prompt treatment, prevention, and control of malarial parasites among HIV-infected patients. In this systematic review and meta-analysis, we estimated the prevalence and characteristics of severe malaria (SM) caused by co-infection with HIV. We included relevant studies that were conducted between the years 1991 and 2018 and reporting on SM. We pooled the prevalence of SM in patients with co-infection, pooled odds ratios of SM in patients with co-infection and Plasmodium mono-infection, and differences in laboratory parameters such as parasite density and leucocyte counts, between co-infected and Plasmodium mono-infected patients. The meta-analysis included 29 studies (1126 SM cases). The pooled prevalence of SM in co-infected patients using the data of 23 studies (SM = 795 cases, all co-infection cases = 2534 cases) was 43.0% (95% confidence interval [CI] 31.0–56.0%; I2, 98.0%). Overall, the odds of SM from 18 studies were pooled. The odds of SM were significantly higher in co-infected patients than in Plasmodium mono-infected patients (OR 2.41; 95% CI 1.43–4.08; I2 = 85%; P = 0.001) and also significantly higher in children (OR 9.69; 95% CI 5.14–18.3; I2, 0%; P < 0.0001; two studies) than in adults (OR 2.68; 95% CI 1.52–4.73; I2, 79.0%; P = 0.0007; 12 studies). Co-infected patients with SM had a higher parasite density than those with Plasmodium mono-infection when the data of seven studies were analysed (SMD, 1.25; 95% CI 0.14–2.36; I2, 98.0%; P = 0.03) and higher leukocyte counts when the data of four studies were analysed (MD, 1570 cells/µL; 95% CI 850–2300 cells/µL; I2, 21.0%; P < 0.0001). Thus, the prevalence of SM among patients co-infected with Plasmodium spp. and HIV is high. Because co-infections could lead to SM, patients with Plasmodium spp. and HIV co-infection should be identified and treated to reduce the prevalence of SM and the number of deaths. [ABSTRACT FROM AUTHOR]

The occurrence of potentially pathogenic free-living amoebae (FLA) in natural hot springs is considered a public health concern. FLAs are known to cause serious health outcomes to a wide spectrum of mammalian hosts. The present study aimed to provide the distribution of isolated cases of FLAs in hot springs through a systematic review process of available published articles online. Relevant studies are published between January 2010 and January 2020 involving the isolation of Naegleria spp., Acanthamoeba spp., Balamuthia spp., Sappinia spp., and Vermamoeba spp. in natural hot springs in the United States, South America, North America, Europe, Asia, and Africa. Articles were identified through a search of PubMed and Google Scholar databases. Out of 94 articles screened, a total of 20 articles are included in the study with consideration of established inclusion and exclusion criteria. The most common FLAs isolated in hot springs are Acanthamoeba spp. (134; 48.5%) and Naegleria spp. (127; 46.0%). Other FLAs isolated in hot springs include Balamuthia spp. (2; 0.7%) and Vermamoeba spp. (13; 4.7%). FLA in hot springs used for recreational and medical purposes is a potential source of infection. It is recommended that strict surveillance and maintenance of hot springs be implemented to prevent potential future infection. [ABSTRACT FROM AUTHOR]

Polymerase chain reaction (PCR) using deoxyribonucleic acid (DNA) extracted from dried blood spots (DBS) provides a fast, inexpensive, and convenient method for large-scale epidemiological studies. This study compared the performance of PCR between DNA extracted from DBS and DNA obtained from whole blood for detecting malarial parasites. Primary studies assessing the diagnostic performance of PCR using DNA extracted from DBS and whole blood for detecting malarial parasites were obtained from the ISI Web of Science, Scopus, and PubMed databases. Odds ratios (ORs) and 95% confidence intervals (CIs) were plotted in forest plots using Review Manager version 5.3. Statistical analysis was performed via random-effects meta-analysis. Data heterogeneity was assessed using the I2 statistic. Of the 904 studies retrieved from the databases, seven were included in this study. The pooled meta-analysis demonstrated no significant difference in the comparative performance of PCR for detecting malaria parasites between DNA extracted from DBS and that extracted from whole blood (OR 0.85; 95% CI 0.62–1.16; I2 = 78%). However, subgroup analysis demonstrated that PCR using DNA extracted from DBS was less accurate in detecting Plasmodium vivax than that using DNA extracted from whole blood (OR = 0.85; 95% CI 0.77–0.94). In conclusion, a significant difference in detecting P. vivax was observed between PCR using DNA extracted from DBS and that using DNA extracted from whole blood. Therefore, P. vivax in endemic areas should be identified and detected with care with PCR using DNA obtained from DBS which potentially leads to a negative result. Further studies are required to investigate the performance of PCR using DBS for detecting P. vivax and other malarial parasites to provide data in research and routine surveillance of malaria, especially with renewed efforts towards the eradication of the disease. [ABSTRACT FROM AUTHOR]

Plasmodium ovale is a benign tertian malaria parasite that morphologically resembles Plasmodium vivax. P. ovale also shares similar tertian periodicity and can cause relapse in patients without a radical cure, making it easily misidentified as P. vivax in routine diagnosis. Therefore, its prevalence might be underreported worldwide. The present study aimed to quantify the prevalence of P. ovale misidentified as P. vivax malaria using data from studies reporting confirmed P. ovale cases by molecular methods. Studies reporting the misidentification of P. ovale as P. vivax malaria were identified from three databases, MEDLINE, Web of Science, and Scopus, without language restrictions, but the publication date was restricted to 1993 and 2020. The quality of the included studies was assessed using the Quality Assessment of Diagnostic Accuracy Studies (QUADAS). The random-effects model was used to estimate the pooled prevalence of the misidentification of P. ovale as P. vivax malaria by the microscopic method when compared to those with the reference polymerase chain reaction method. Subgroup analysis of participants was also performed to demonstrate the difference between imported and indigenous P. ovale cases. The heterogeneity of the included studies was assessed using Cochran's Q and I2 statistics. Publication bias across the included studies was assessed using the funnel plot and Egger's test, and if required, contour-enhanced funnel plots were used to identify the source(s) of funnel plot asymmetry. Of 641 articles retrieved from databases, 22 articles met the eligibility criteria and were included in the present study. Of the 8,297 malaria-positive cases identified by the PCR method, 453 P. ovale cases were confirmed. The pooled prevalence of misidentification of P. ovale as P. vivax malaria by the microscopic method was 11% (95% CI: 7–14%, I2: 25.46%). Subgroup analysis of the participants demonstrated a higher prevalence of misidentification in indigenous cases (13%, 95% CI: 6–21%, I2: 27.8%) than in imported cases (10%, 95% CI: 6–14%, I2: 24.1%). The pooled prevalence of misidentification of P. vivax as P. ovale malaria by the microscopic method was 1%, without heterogeneity (95% CI: 0–3%, I2: 16.8%). PCR was more sensitive in identifying P. ovale cases than the microscopic method (p < 0.00001, OR: 2.76, 95% CI: 1.83–4.15, I2: 65%). Subgroup analysis of participants demonstrated the better performance of PCR in detecting P. ovale malaria in indigenous cases (p: 0.0009, OR: 6.92, 95% CI: 2.21–21.7%, I2: 68%) than in imported cases (p: 0.0004, OR: 2.15, 95% CI: 1.41–3.29%, I2: 63%). P. ovale infections misidentified as P. vivax malaria by the microscopic method were frequent and led to underreported P. ovale cases. The molecular identification of P. ovale malaria in endemic areas is needed because a higher rate of P. ovale misidentification was found in endemic or indigenous cases than in imported cases. In addition, updated courses, enhanced training, and refreshers for microscopic examinations, particularly for P. ovale identification, are necessary to improve the microscopic identification of Plasmodium species in rural health centres where PCR is unavailable. [ABSTRACT FROM AUTHOR]

Malaria rapid diagnostic tests (RDTs) are widely used to detect malaria parasites among patients who suspected malaria infections in malaria-endemic areas where microscopy is unavailable. Nevertheless, little is known about the performance of RDTs in detecting Plasmodium mixed infections. The present study aimed to evaluate the discordant results between RDTs and microscopy/polymerase chain reaction (PCR) in detecting Plasmodium mixed infections. The PubMed (MEDLINE), Web of Science, and Scopus databases were systematically reviewed to identify related studies that reported the performance of RDTs in detecting Plasmodium mixed infections. Studies were grouped according to the different RDT types including RDT type 2 (pf-HRP2/pan-aldolase), RDT type 3 (pf-HRP2/pan-pLDH), RDT type 4 (Pf-LDH/pan-pLDH), RDT type 5 (Pf/Pv-pLDH), and RDT type 6 (pf-HRP2/Pv-pLDH) for subgroup analysis. The estimates of the different proportions in each analysis group that were visually summarized in a forest plot showed the odds ratio (OR) and 95% confidence interval (CI). Plots were drawn using RevMan (version 5.3; Cochrane Community). Twenty-eight studies were included in the present study. Overall, the meta-analysis showed that RDTs could detect a significantly higher proportion of Plasmodium mixed infections than microscopy (p = 0.0007, OR = 3.33, 95% CI 1.66–6.68). Subgroup analysis demonstrated that only RDTs targeting Pf-specific histidine-rich protein 2 (HRP2)/pan-specific lactate dehydrogenase (LDH) could detect a significantly higher proportion of Plasmodium mixed infections than microscopy (p = 0.004, OR = 8.46, 95% CI 2.75–26.1). The subgroup analysis between RDTs and PCR methods demonstrated that RDTs targeting Pf-specific HRP2/Pv-specific LDH could detect a significantly lower proportion of Plasmodium mixed infections than PCR methods (p = 0.0005, OR = 0.42, 95% CI 0.26–0.68). This is the first study to summarize the discordant results between RDTs and microscopy/PCR in detecting Plasmodium mixed infections. Malaria RDTs targeting Pf-HRP2/pan-pLDH could detect a higher proportion of Plasmodium mixed infections than microscopy, while RDTs targeting Pf-HRP2/Pv-specific LDH could detect a lower proportion of Plasmodium mixed infections than PCR methods. The results of this study will support the selection and careful interpretations of RDTs for a better diagnosis of Plasmodium mixed-species infections and appropriate treatment of malaria patients in endemic and non-endemic settings. [ABSTRACT FROM AUTHOR]

Mixed Plasmodium malaria infections can lead to severe malaria. This systematic review and meta-analysis aimed to explore the prevalence of severe mixed Plasmodium malaria infection and to compare it with the prevalence of severe P. falciparum malaria mono-infection across the included studies. Original English-language research articles from PubMed, Scopus, and ISI Web of Science were identified and screened. Articles reporting the number of mixed infections and the number of severe mixed infections were used to determine the main outcome of this study, while the number of P. falciparum infections and the number of severe P. falciparum infections were used to determine the secondary outcome of this study. For the main outcome, the pooled prevalence and 95% confidence interval (CI) of severe mixed infections was analysed using STATA software version 15.0 (Stata Corp, College Station, TX, USA). For the secondary outcome, the rate of severe mixed infections compared to severe P. falciparum infections was analysed using the meta-analysis approach, and summary odds ratios (ORs) and 95% CIs were calculated. Random-effects models were used to produce the summary ORs. The Mantel–Haenszel method and calculated I2 were also reported to test whether there was heterogeneity among the included studies. Publication bias was also assessed using funnel plots. The meta-analysis of secondary outcomes was conducted using Review Manager 5.3 software (Cochrane Community). A total of 894,561 malaria patients were reported in all 16 included studies. Overall, a pooled analysis showed that 9% (2,006/35,768, 95% CI 7.0–12.0%) of patients with mixed Plasmodium infection had severe mixed infection. A meta-analysis of 14 studies demonstrated that patients with mixed Plasmodium infection (1,999/35,755) and patients with P. falciparum malaria (9,249/294,397) had an equal risk of developing severe malaria (OR 0.93, 95% CI 0.59–1.44). Both mixed infection and P. falciparum mono-infection showed a similar trend of complications in which severe anaemia, pulmonary failure, and renal impairment were the three most common complications found. However, patients with mixed infection had a higher proportion of severe anaemia and pulmonary complications than those with P. falciparum infection. Moreover, patients with mixed infection had a higher proportion of multiple organ failure than those with P. falciparum mono-infection. Mixed Plasmodium spp. infections were common but often unrecognized or underestimated, leading to severe complications among these malaria patients. Therefore, in routine clinical laboratories, using an accurate combination of diagnostic procedures to identify suspected patients with mixed infections is crucial for therapeutic decisions, prompt treatment, and effective patient management. [ABSTRACT FROM AUTHOR]