Molecular Surveillance for Imported Antimicrobial Resistant Plasmodium falciparum, Ontario, Canada.

Single-nucleotide polymorphisms at several loci have been correlated with Plasmodium falciparum drug resistance. We examined the prevalence of resistance markers in P. falciparum from imported malaria cases in Canada during 3 time periods, 2008-2009, 2013-2014, and 2017-2018. We evaluated single-nucleotide polymorphisms at atpase6 (pfATPase6), pfcrt (chloroquine resistance transporter), cytb (cytochrome b), dhfr (dihydrofolate reductase), dhps (dihydropteroate synthetase), mdr1 (multidrug resistance protein) and mdr1 copy number, and kelch13 (kelch protein gene on chromosome 13). Over time, we observed increasing mutant genotypes for dhfr S108N and dhps A613T and decreasing mutant genotypes for mdr1 N86Y, D1246Y, pfcrt K76T, and pfcrt 74-75; we identified no kelch13 mutations. We observed fewer mutations indicative of chloroquine resistance over time, which may reflect reduced chloroquine pressure in specimens from travelers to Africa. Mutations conferring proguanil resistance increased over time. Minor genotypes confirm the heterogeneous nature of infection and may affect treatment success.

Trichinella Nativa Outbreak With Rare Thrombotic Complications Associated With Meat From a Black Bear Hunted in Northern Ontario.

BACKGROUND: Although trichinellosis is known to cause thrombotic disease, serious thrombotic events are rare and have not been previously associated with Trichinella nativa infection. METHODS: Patient interviews and medical chart reviews were conducted on 10 men who became ill following consumption of a common source of black bear meat. Trichinella serology on patient sera as well as polymerase chain reaction (PCR) and larval identification of the meat samples was conducted. RESULTS: All 10 exposed individuals developed an acute illness clinically compatible with trichinellosis, characterized by fever, abdominal pain, and diarrhea, along with eosinophilia ranging from 0.9 × 109/L to 6.1 × 109/L. Within 2 weeks of the diarrheal illness, systemic symptoms developed in all exposed individuals characterized by fever, myalgia, periorbital edema, and fatigue. ST-elevation myocardial infarction and sinus venous tract thrombosis occurred as a complication of trichinellosis in 2 patients. Acute serology was nonreactive in all patients, though convalescent serology was reactive in 6 of 8 (75%) patients for whom sera was available. Multiplex PCR identified T. nativa from the bear meat, and was corroborated by microscopic larval identification. CONCLUSIONS: We report a 100% attack rate of T. nativa from bear meat among those who were exposed, and demonstrate that this species can cause serious thrombotic complications of trichinellosis in humans. Education of hunters and the public regarding the importance of proper preparation of wild game prior to ingestion is warranted.

Parasitological correlates of Plasmodium ovale curtisi and Plasmodium ovale wallikeri infection.

BACKGROUND: Malaria, due to Plasmodium ovale, can be challenging to diagnose due to clinically mild disease and low parasite burden. Two genetically distinct sub-species of P. ovale exist: Plasmodium ovale curtisi (classic) and Plasmodium ovale wallikeri (variant). It is presently unknown if the sub-species causing infection affects performance of malaria diagnostic tests. The aim of this work was to understand how the genetically distinct sub-species, P. o. curtisi and P. o. wallikeri, affect malaria diagnostic tests. METHODS: Plasmodium ovale-positive whole blood specimens were sub-speciated by PCR and sequencing of 18S rRNA and dhfr-ts. Parasitaemia, morphology, pan-aldolase positivity, 18S copy number, and dhfr-ts sequences were compared between sub-species. RESULTS: From 2006 to 2015, 49 P. ovale isolates were identified, of which 22 were P. o. curtisi and 27 P. o. wallikeri; 80% were identified in the last five years, and 88% were acquired in West Africa. Sub-species did not differ by parasitaemia, 18S copy number, or pan-aldolase positivity. Lack of Schüffner's stippling was over-represented among P. o. wallikeri isolates (p = 0.02). Several nucleotide polymorphisms between the sub-species were observed, but they do not occur at sites believed to relate to antifolate binding. CONCLUSIONS: Plasmodium ovale is increasing among travellers to West Africa, although sub-species do not differ significantly by parasitologic features such as parasitaemia. Absence of Schüffner's stippling may be a feature specific to P. o. wallikeri and is a novel finding.

Evaluation of Ebola virus inactivation procedures for Plasmodium falciparum malaria diagnostics.

Plasmodium falciparum malaria is highly endemic in the three most affected countries in the current epidemic of Ebola virus disease (EVD) in West Africa. As EVD and malaria are clinically indistinguishable, both remain part of the differential diagnosis of ill travelers from returning from areas of EVD transmission. We compared the performances of a rapid diagnostic test (BinaxNOW) and real-time PCR with P. falciparum-positive specimens before and after heat and Triton X-100 inactivation, and we documented no loss of sensitivity.

Fever in a visitor to Canada: a case of mistaken identity.

We report a case of babesiosis in a traveler from India who was diagnosed with malaria on the basis of blood smears. Pan-Plasmodium PCR was positive, though species-specific assays were negative. Reexamination of blood smears and Babesia-specific PCR confirmed babesiosis. We highlight the overlapping clinical and diagnostic features of malaria and babesiosis and the potential cross-reactivity of Plasmodium primers in cases of babesiosis.

Reevaluation of an Acanthamoeba Molecular Diagnostic Algorithm following an Atypical Case of Amoebic Keratitis.

Amoebic keratitis (AK) is a potentially blinding infection, the prompt diagnosis of which is essential for limiting ocular morbidity. We undertook a quality improvement initiative with respect to the molecular detection of acanthamoebae in our laboratory because of an unusual case of discordance. Nine ATCC strains of Acanthamoeba and 40 delinked, biobanked, surplus corneal scraping specimens were analyzed for the presence of acanthamoebae with four separate real-time PCR assays. The assay used by the Free-Living and Intestinal Amebas Laboratory of the CDC was considered the reference standard, and the performance characteristics of each individual assay and pairs of assays were calculated. Outcome measures were sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV). Of 49 included specimens, 14 (28.6%) were positive by the gold standard assay, and 35 (71.4%) were negative. The sensitivities of the individual assays ranged from 64.3% to 92.9%, compared to the gold standard, while the specificities ranged from 88.6% to 91.4%. The PPVs and NPVs ranged from 69.2% to 78.6% and from 86.1% to 96.9%, respectively. Combinations of assay pairs led to improved performance, with sensitivities ranging from 92.9% to 100% and specificities ranging from 97.1% to 100%. ATCC and clinical strains of Acanthamoeba that failed to be detected by certain individual assays included Acanthamoeba castellanii, Acanthamoeba culbertsoni, and Acanthamoeba lenticulata. For three clinical specimens, false negativity of the gold standard assay could not be excluded. Molecular diagnostic approaches, especially combinations of highly sensitive and specific assays, offer a reasonably performing, operator-independent, rapid strategy for the detection of acanthamoebae in clinical specimens and are likely to be more practical than either culture or direct microscopic detection.

Survival analysis of diagnostic assays in Plasmodium falciparum malaria.

BACKGROUND: Rapid diagnostic tests (RDT) and real-time PCR (qPCR) assays are sensitive for diagnosing malaria, but because they detect antigen and DNA, respectively, positivity may not reflect active infection. Performance characteristics of RDT and qPCR in Plasmodium falciparum positive specimens were evaluated over time to elucidate duration of positivity following conversion to microscopy negative. METHODS: Specimens from patients with at least one specimen that was positive for P. falciparum by microscopy, and at least one specimen that was negative for P. falciparum within a 1-month period were identified. Survival distributions of the diagnostic tests over time were compared. Performance characteristics for each test were calculated. RESULTS: Ninety specimens were included, with 48 initially positive for P. falciparum, and 42 subsequently negative. Of 42 specimens that converted to microscopy-negative following an initial positive, 26 (61.9 %) and 41 (97.6 %) were positive by qPCR and RDT, respectively. Survival curves of microscopy versus qPCR, as well as microscopy vs RDT differed significantly (p = 0.0002 and p < 0.0001, respectively). Compared to microscopy, sensitivity of qPCR was 100.0 % (95 % CI 90.8-100.0 %), and that of RDT was 100.0 % (95 % CI 90.8-100.0 %). CONCLUSIONS: Due to slow clearance of circulating antigen and DNA from bloodstream, RDT and qPCR have low positive predictive value for clinically relevant asexual parasitaemia in post-treatment specimens. Thus, microscopy remains the only available malaria diagnostic that can reliably distinguish true asexual parasitaemia from prolonged clearance of antigen and nucleic acid in a convalescing patient.

An elderly long-term care resident with crusted scabies.

Crusted scabies is a highly contagious form of scabies. Altered immune response, nutritional deficiencies and modified host response are all risk factors for crusted scabies. The authors report a case involving a patient found to have a chronic maculopapular, erythematous rash with large hyperkeratotic, white and grey plaques on the soles of both feet. An ultimate diagnosis of crusted scabies was reached after a delay in diagnosis suspected to be caused by the similarity in appearance to more common skin conditions such as psoriasis. After topical permethrin was unsuccessful, intermittent dosing of oral ivermectin resulted in a rapid reduction in cutaneous plaques.

La gale hyperkératosique est une forme de gale extrêmement contagieuse. Une détérioration de la réponse immunitaire, des carences nutritionnelles et une réponse modifiée de l'hôte en sont toutes des facteurs de risque. Les auteurs rendent compte d'un patient présentant une éruption maculopapuleuse et érythémateuse chronique comportant de grosses plaques hyperkératosiques blanches et grises sur la plante des deux pieds. Le diagnostic de gale hyperkératosique a été posé après un retard de diagnostic qu'on croit attribuable à sa similarité d'aspect avec des troubles cutanés plus courants, tels que le psoriasis. Après avoir essayé la perméthrine topique sans succès, une posologie intermittente d'ivermectine par voie orale a suscité la réduction rapide des plaques cutanées.

Sequence-based optimization of a quantitative real-time PCR assay for detection of Plasmodium ovale and Plasmodium malariae.

Although microscopic examination of Giemsa-stained blood smears remains the gold standard for the diagnosis of malaria, molecular detection using PCR is becoming increasingly popular. Due to discrepant PCR and microscopy results, we aimed to optimize our detection assays for Plasmodium malariae and Plasmodium ovale by sequencing the 18S rRNA region and developing a new primer and probe set for real-time quantitative PCR (qPCR). Clinical specimens positive for P. malariae (n = 15) or P. ovale (n = 33) underwent amplification and sequencing of the 18S rRNA region. Based on sequence discrepancies between our current primer/probe and clinical isolates, degenerate P. ovale primer and probe were developed to determine if their performance characteristics improved. The reference (gold) standard was microscopy. No 18S sequence heterogeneity was observed among the P. malariae isolates, and the sensitivity and specificity of our current P. malariae qPCR assay were both 100%. Compared to microscopy, the sensitivity and specificity of our current P. ovale qPCR assay were 72.7% and 100%, respectively. Five single nucleotide polymorphisms (SNPs) were identified in P. ovale. The sensitivity of the new P. ovale assay increased to 100% with 100% specificity. We therefore improved the performance characteristics of our P. ovale molecular detection assay through the development of a degenerate primer and probe set which accommodates 18S SNPs among the 2 subspecies of P. ovale. Given the suboptimal sensitivity of rapid diagnostic tests for non-falciparum malaria and the typically low parasitemia of P. malariae and P. ovale, a well-performing confirmatory molecular assay is imperative for clinical laboratories.

Performance characteristics of diagnostic assays for schistosomiasis in Ontario, Canada.

Introduction: Due to lower intensity of infection and greater intervals from last exposure, parasitologic detection methods for schistosomiasis are poorly sensitive in non-endemic areas, challenging accurate diagnosis. Methods: We evaluated parasitologic versus indirect detection methods for schistosomiasis. We included specimens submitted for Schistosoma serology, and stool for ova and parasite microscopy. Three real-time PCR assays targeting Schistosoma mansoni and S. haematobium were performed. Primary outcome measures were sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV), where both microscopy and serology were the composite reference standard against serum PCR. Results: Of 8168 serum specimens submitted for Schistosoma serology, 638 (7.8%) were reactive and 6705 (82.1%) were non-reactive. Of 156,771 stool specimens submitted for ova and parasite testing, 46 (0.03%) were positive for eggs of S. mansoni. Four (0.5%) urine specimens were positive for eggs of S. haematobium. Combined serum PCRs targeting S. mansoni had a sensitivity and specificity of 27.8% (95% CI = 18.3-39.1%) and 100% (95% CI = 83.9-100%), respectively, with PPV of 100% (95% CI = 100%) and NPV of 26.9% (95% CI = 24.3-29.7%). The one serum sample positive for S. haematobium was also detectable by our S. haematobium PCR. No cross-reactivity was observed for all three PCR assays. Conclusions: Although serology is highly sensitive, parasitologic tests signify active infection, but are limited by low population-level sensitivity, particularly in non-endemic settings. Although serum PCR offered no performance advantage over stool microscopy, its role in diagnostic parasitology should be pursued due to its high-throughput and operator-independent nature.

Surveillance of Amoebic Keratitis-Causing Acanthamoebae for Potential Bacterial Endosymbionts in Ontario, Canada.

Acanthamoeba spp. are the causative pathogens of several infections, including amoebic keratitis (AK), a vision-threatening infection. Acanthamoebae from corneal specimens of patients with AK harbor bacterial endosymbionts, which may increase virulence. We sought to understand the spectrum of bacterial endosymbionts present in clinical isolates of Acanthamoeba spp. identified in our reference parasitology laboratory. Isolates of Acanthamoeba spp. obtained from our biobank of anonymized corneal scrapings were screened for potential endosymbionts by PCR using primer pairs detecting bacteria belonging to orders Chlamydiales, Rickettsiales, or Legionellales and pan16S primers. Three primer pairs specific to the 18s rRNA gene of Acanthamoeba spp. were used for the amplification of Acanthamoeba DNA used for sequencing. Sanger sequencing of all PCR products was performed, followed by BLAST analysis for species identification. We screened 26 clinical isolates of Acanthamoeba spp. for potential endosymbionts. Five isolates (19%) were found to contain bacterial DNA belonging to Legionellales. Three (11%) contained members of the Rickettsiales and Pseudomonas genticulata was detected in a Rickettsia-positive sample. One strain (4%) contained Neochlamydia hartmannellae, a member of the Chlamydiales order. Bacterial endosymbionts are prevalent in clinical strains of Acanthamoeba causing AK isolated from corneal scrapings. The demonstration of these organisms in clinical Acanthamoeba isolates supports a potential exploration of anti-endosymbiont therapeutics as an adjuvant therapy in the treatment of AK.

Multiplex real-time quantitative PCR, microscopy and rapid diagnostic immuno-chromatographic tests for the detection of Plasmodium spp: performance, limit of detection analysis and quality assurance.

BACKGROUND: Accurate laboratory diagnosis of malaria species in returning travelers is paramount in the treatment of this potentially fatal infectious disease. MATERIALS AND METHODS: A total of 466 blood specimens from returning travelers to Africa, Asia, and South/Central America with suspected malaria infection were collected between 2007 and 2009 at the reference public health laboratory. These specimens were assessed by reference microscopy, multipex real-time quantitative polymerase chain reaction (QPCR), and two rapid diagnostic immuno-chromatographic tests (ICT) in a blinded manner. Key clinical laboratory parameters such as limit of detection (LOD) analysis on clinical specimens by parasite stage, inter-reader variability of ICTs, staffing implications, quality assurance and cost analysis were evaluated. RESULTS: QPCR is the most analytically sensitive method (sensitivity 99.41%), followed by CARESTART (sensitivity 88.24%), and BINAXNOW (sensitivity 86.47%) for the diagnosis of malaria in returning travelers when compared to reference microscopy. However, microscopy was unable to specifically identify Plasmodia spp. in 18 out of 170 positive samples by QPCR. Moreover, the 17 samples that were negative by microscopy and positive by QPCR were also positive by ICTs. Quality assurance was achieved for QPCR by exchanging a blinded proficiency panel with another reference laboratory. The Kappa value of inter-reader variability among three readers for BINAXNOW and CARESTART was calculated to be 0.872 and 0.898 respectively. Serial dilution studies demonstrated that the QPCR cycle threshold correlates linearly with parasitemia (R(2) = 0.9746) in a clinically relevant dynamic range and retains a LOD of 11 rDNA copies/microl for P. falciparum, which was several log lower than reference microscopy and ICTs. LOD for QPCR is affected not only by parasitemia but the parasite stage distribution of each clinical specimen. QPCR was approximately 6-fold more costly than reference microscopy. DISCUSSION: These data suggest that multiplex QPCR although more costly confers a significant diagnostic advantage in terms of LOD compared to reference microscopy and ICTs for all four species. Quality assurance of QPCR is essential to the maintenance of proficiency in the clinical laboratory. ICTs showed good concordance between readers however lacked sensitivity for non-falciparum species due to antigenic differences and low parasitemia. CONCLUSION: Multiplex QPCR but not ICTs is an essential adjunct to microscopy in the reference laboratory detection of malaria species specifically due to the superior LOD. ICTs are better suited to the non-reference laboratory where lower specimen volumes challenge microscopy proficiency in the non-endemic setting.

Occurrence and distribution of Ambylomma americanum as determined by passive surveillance in Ontario, Canada (1999-2016).

The lone star tick, Amblyomma americanum, is spreading northward from its historical stronghold in the southeastern United States. As a vector and biting pest, public and veterinary health officials must remain vigilant of the lone star tick's expanding range. We use ticks submitted to Public Health Ontario Laboratory (1999-2016) to describe the spatial and temporal dynamics of A. americanum in Ontario, Canada, as well as submitter demographics. We identified 847 A. americanum submissions during the surveillance period, with 773 (91.3%) non-travel-related and 74 (8.7%) travel-related submissions. Annual A. americanum submissions increased over the surveillance period. Approximately 91% of non-travel-related submissions were adult ticks and 9% were nymphs. The highest submission rates were from individuals living in the Eastern and South West regions of the province. Adult specimens were primarily submitted from May through July and nymphs from March through September. Higher numbers of submissions were from young children (<10 years) and older adults (55-74 years), with equal proportions of male and female submitters. The majority of travel-related submissions were from travellers returning from the southeastern United States (i.e., Florida, North Carolina, South Carolina, Tennessee, Texas). Amblyomma americanum distribution is scattered in Ontario and submissions are likely the consequence of ongoing detection of adventive specimens. Further tick dragging is required to confirm the presence of established lone star tick populations in the province. Given the relatively rapid expansion of blacklegged ticks, Ixodes scapularis, populations in Ontario, we expect climate change to facilitate the range of expansion of A. americanum into the province. We propose an algorithm for identifying A. americanum-risk areas, which will aid public and veterinary health officials when assessing the risks posed by lone star ticks.

Development and validation of a real-time PCR assay for the detection of clinical acanthamoebae.

BACKGROUND: Suboptimal agreement between molecular assays for the detection of Acanthamoeba spp. in clinical specimens has been demonstrated, and poor assay sensitivity directly imperils the vision of those affected by amoebic keratitis (AK) through delayed diagnosis. We sought to develop and validate a single Taqman real time PCR assay targeting the Acanthamoeba 18S rRNA gene that could be used to enhance sensitivity and specificity when paired with reference assays. METHODS: Biobanked DNA from surplus delinked AK clinical specimens and 10 ATCC strains of Acanthamoeba was extracted. Sequence alignment of 66 18S rRNA regions from 12 species of Acanthamoeba known to cause keratitis informed design of a new TaqMan primer set. Performance of the new assay was compared to the 2 assays used currently in our laboratory. RESULTS: Among 24 Acanthamoeba-positive and 83 negative specimens by the CDC reference standard, performance characteristics of the newly designed primer set were as follows: sensitivity 100%, specificity 94%, PPV 82.8%, and NPV 100%. Compared to culture, sensitivity of the new primer set was 100%, and specificity 96%. No cross-reactivity of the primer set to non-acanthamoebae, including Balamuthia and Naegleria, was found. CONCLUSIONS: We have validated a real time PCR assay for the diagnosis of AK, and in doing so, have overcome important barriers to rapid and sensitive detection of acanthamoebae, including limited sensitivity and specificity of commonly used assays.

Prevalence of Rickettsia species in Dermacentor variabilis ticks from Ontario, Canada.

Relatively little is known about the prevalence of rickettsial species in Dermacentor ticks in eastern Canada. In this study, Dermacentor ticks from the province of Ontario, Canada, were tested for the presence of spotted fever group rickettsial (SFGR) species, Coxiella burnetii and Francisella tularensis. Rickettsia rickettsii was not detected in any ticks tested, but R. montanensis was detected at a prevalence of 2.2% in D. variabilis (17/778). Two other SFGR species, R. parkeri and Candidatus R. andeanae, were detected individually in 2 Amblyomma maculatum ticks. Rickettsia peacockii, a non-pathogenic endosymbiont, was detected in two D. andersonii ticks. Given the highly abundant nature of D. variabilis, surveillance for human pathogens in this species of tick has important public health implications, but the lack of detection of known human pathogens indicates a low risk of infection via this tick species in Ontario. However, the detection of R. parkeri in an adventive A. maculatum tick indicates that health care providers should be aware of the possibility of spotted fever rickettsioses in individuals with a history of travel outside of Ontario and symptoms compatible with a spotted fever rickettsiosis. Coxiella burnetii and Francisella tularensis, human pathogens also potentially transmitted by D. variabilis, were not detected in a subset of the ticks.

Strongyloidiasis in Ontario: Performance of diagnostic tests over a 14-month period.

BACKGROUND: We evaluated the performance of stool microscopy, serology, and real time PCR (qPCR) for the diagnosis of strongyloidiasis at our reference laboratory. METHODS: Using a convenience sample of specimens submitted between April 1, 2014 and May 31, 2015, positivity rates and performance characteristics were calculated. RESULTS: During the enrolment period, 17,933 stool specimens were examined for O&P, 14 of which were positive for Strongyloides larvae. For stool specimens serially positive for larvae, mean duration of larval shedding was 12.7 days following the initial positive specimen, while for sputum and urine, it was 12 and 2 days, respectively. During the enrolment period, 3258 specimens were processed for Strongyloides serology, 200 of which were reactive (6.1%), 210 indeterminate (6.5%), and 2848 non-reactive (87.4%). qPCR was positive in 11 of 12 (91.7%) stool specimens containing larvae, and negative in all stool specimens without larvae by microscopy. There was no cross-reactivity of Strongyloides-specific qPCR to other stool protozoa or helminths. CONCLUSIONS: In the absence of immunosuppression, larval burden in strongyloidiasis is low, limiting the utility of microscopy, and favoring serologic testing. However, false negative serology can occur in those with hyperinfection necessitating a combined diagnostic approach. qPCR was insufficiently sensitive to replace microscopy for detection of larvae.

Correlating quantitative real-time PCR to rapid diagnostic test and RNA transcript expression in isolated gametocytemia and asexual parasitemia of Plasmodium falciparum malaria.

BACKGROUND: At present, only microscopic examination of stained thick and thin blood smears for malaria can differentiate clinically relevant asexual parasitemia from clinically irrelevant isolated gametocytemia. Microscopy is time consuming, labour intensive, and requires significant technical expertise to perform. Simple and rapid tests that can distinguish asexual from isolated sexual parasitemia are needed. METHODS: To determine if parasitemia and cycle threshold (CT) values on Plasmodium genus and P. falciparum-specific quantitative polymerase chain reaction (qPCR) assays correlate to positivity of rapid diagnostic test (RDT), and 18S rRNA gene copy number, we analyzed blood samples from Ontario patients with isolated P. falciparum gametocytemia or asexual stages. RNA transcripts were evaluated to determine whether there is correlation of expression to different life cycle stages of P. falciparum. RESULTS: 45 specimens containing isolated P. falciparum gametocytes, and 40 specimens containing isolated asexual stages by microscopy were identified and analyzed. By RDT, 40 of 45 (88.9 %) isolated gametocytemia specimens and 40 of 40 (100 %) asexual-stage specimens were positive for Plasmodium falciparum-specific histidine rich protein-2 (HRP-2). Fourteen of 45 (31.1 %) isolated gametocytemia specimens, and 36 of 40 (90 %) asexual-stage specimens were positive for Plasmodium genus aldolase T2 band. Positivity of the aldolase T2 band was associated with lower mean Plasmodium genus and P. falciparum-specific CT values, and to higher mean 18S rRNA gene copy by qPCR for both isolated gametocytemia and asexual-stage specimens. There was also a negative correlation of asexual parasitemia to both CT values, and positive correlation to 18S rRNA gene copy number. Analysis of asexual stage-specific erythrocyte binding antigen (eba-175) transcripts on 25 isolated gametocytemia and 20 asexual-stage specimens gave a positive predictive value of 62.5 % and negative predictive value of 100 % for asexual parasitemia. Thus, an absence of eba-175 transcripts excluded the presence of asexual (clinically relevant) parasitemia. CONCLUSIONS: Positivity of the aldolase T2 band of BinaxNow RDT correlated to higher parasite load in both isolated gametocytemia and asexual-stage specimens. Asexual stage-specific eba-175 RNA transcript expression provided reasonable negative predictive value for exclusion of asexual parasitemia in clinical samples, but was present in both isolated gametocytemia and asexual stage specimens.

Population-based passive tick surveillance and detection of expanding foci of blacklegged ticks Ixodes scapularis and the Lyme disease agent Borrelia burgdorferi in Ontario, Canada.

We identified ticks submitted by the public from 2008 through 2012 in Ontario, Canada, and tested blacklegged ticks Ixodes scapularis for Borrelia burgdorferi and Anaplasma phagocytophilum. Among the 18 species of ticks identified, I. scapularis, Dermacentor variabilis, Ixodes cookei and Amblyomma americanum represented 98.1% of the 14,369 ticks submitted. Rates of blacklegged tick submission per 100,000 population were highest in Ontario's Eastern region; D. variabilis in Central West and Eastern regions; I. cookei in Eastern and South West regions; and A. americanum had a scattered distribution. Rates of blacklegged tick submission per 100,000 population were highest from children (0-9 years old) and older adults (55-74 years old). In two health units in the Eastern region (i.e., Leeds, Grenville & Lanark District and Kingston-Frontenac and Lennox & Addington), the rate of submission for engorged and B. burgdorferi-positive blacklegged ticks was 47× higher than the rest of Ontario. Rate of spread for blacklegged ticks was relatively faster and across a larger geographic area along the northern shore of Lake Ontario/St. Lawrence River, compared with slower spread from isolated populations along the northern shore of Lake Erie. The infection prevalence of B. burgdorferi in blacklegged ticks increased in Ontario over the study period from 8.4% in 2008 to 19.1% in 2012. The prevalence of B. burgdorferi-positive blacklegged ticks increased yearly during the surveillance period and, while increases were not uniform across all regions, increases were greatest in the Central West region, followed by Eastern and South West regions. The overall infection prevalence of A. phagocytophilum in blacklegged ticks was 0.3%. This study provides essential information on ticks of medical importance in Ontario, and identifies demographic and geographic areas for focused public education on the prevention of tick bites and tick-borne diseases.