Publications by authors named "S Hendrickx"

68 Publications

Percutaneous tracheostomy for long-term ventilated COVID-19-patients: rationale and first clinical-safe for all-experience.

Anaesthesiol Intensive Ther 2020;52(5):366-372

Department of Intensive Care, Universitair Ziekenhuis Brussel, Jette, Brussels, Belgium.

Introduction: COVID-19 infection has resulted in thousands of critically ill patients admitted to ICUs and treated with mechanical ventilation. Percutaneous tracheostomy is a well-known technique utilised as a strategy to wean critically ill patients from mechanical ventilation. Worldwide differences exist in terms of methods, operators, and settings, and questions remain regarding timing and indications. If tracheostomy is to be performed in COVID-19 patients, a safe environment is needed for optimal care.

Material And Methods: We present a guidewire dilating forceps tracheostomy procedure in COVID-19 patients that was optimised including apnoea-moments, protective clothing, checklists, and clear protocols. We performed a retrospective analysis of the outcome after tracheostomy in COVID-19 patients between March 2020 and May 2020.

Results: The follow-up of the first 16 patients, median age 62 years, revealed a median intubation time until tracheostomy of 18 days and median cannulation time of 20 days. The overall perioperative complication rate and complication rate while cannulated was 19%, mainly superficial bleeding. None of the healthcare providers involved in performing the procedure developed any symptoms of the disease.

Conclusions: This COVID-19-centred strategy based on flexibility, preparation, and cooperation between healthcare providers with different backgrounds facilitated percutaneous tracheostomy in COVID-19 patients without an increase in the overall complication rate or evidence of risk to healthcare providers. Our findings provide initial evidence that tracheostomy can be performed safely as a standard of care for COVID-19 patients requiring prolonged mechanical ventilation as was standard practice in ICU patients prior to the COVID-19 pandemic to promote ventilator weaning and patient recovery.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.5114/ait.2020.101216DOI Listing
December 2020

Correction to: Preparation and Characterization of Nanostructured Lipid Carriers for Improved Topical Drug Delivery: Evaluation in Cutaneous Leishmaniasis and Vaginal Candidiasis Animal Models.

AAPS PharmSciTech 2020 Oct 8;21(7):275. Epub 2020 Oct 8.

Department of Pharmacy, Quaid-i-Azam University, Islamabad, 45320, Pakistan.

In the published manuscript, co-author Sarah Hendrickx name was misspelled and co-author Guy Caljon's last and first names were inadvertently switched.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1208/s12249-020-01819-5DOI Listing
October 2020

Sand Fly Studies Predict Transmission Potential of Drug-resistant Leishmania.

Trends Parasitol 2020 09 23;36(9):785-795. Epub 2020 Jul 23.

Laboratory of Microbiology, Parasitology and Hygiene (LMPH), University of Antwerp, Universiteitsplein 1, B-2610 Wilrijk, Belgium. Electronic address:

Leishmania parasites have the capacity to rapidly adapt to changing environments in their digenetic life cycle which alternates between a vertebrate and an invertebrate host. Emergence of resistance following drug exposure can evoke phenotypic alterations that affect several aspects of parasite fitness in both hosts. Current studies of the impact of resistance are mostly limited to interactions with the mammalian host and characterization of in vitro parasite growth and differentiation. Development in the vector and transmission capacity have been largely ignored. This review reflects on the impact of drug resistance on its spreading potential with specific focus on the use of the sand fly infection model to evaluate parasite development in the vector and the ensuing transmission potential of drug-resistant phenotypes.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.pt.2020.06.006DOI Listing
September 2020

Antileishmanial Aminopyrazoles: Studies into Mechanisms and Stability of Experimental Drug Resistance.

Antimicrob Agents Chemother 2020 08 20;64(9). Epub 2020 Aug 20.

Laboratory of Microbiology, Parasitology and Hygiene (LMPH), University of Antwerp, Antwerp, Belgium

Current antileishmanial treatment is hampered by limitations, such as drug toxicity and the risk of treatment failure, which may be related to parasitic drug resistance. Given the urgent need for novel drugs, the Drugs for Neglected Diseases (DND) has undertaken a drug discovery program, which has resulted in the identification of aminopyrazoles, a highly promising antileishmanial chemical series. Multiple experiments have been performed to anticipate the propensity for resistance development. Resistance selection was performed by successive exposure of promastigotes () and intracellular amastigotes (both and in golden Syrian hamsters). The stability of the resistant phenotypes was assessed after passage in mice and sandflies. Whole-genome sequencing (WGS) was performed to identify mutated genes, copy number variations (CNVs), and somy changes. The potential role of efflux pumps (the MDR and MRP efflux pumps) in the development of resistance was assessed by coincubation of aminopyrazoles with specific efflux pump inhibitors (verapamil, cyclosporine, and probenecid). Repeated drug exposure of amastigotes did not result in the emergence of drug resistance either or Selection at the promastigote stage, however, was able to select for parasites with reduced susceptibility (resistance index, 5.8 to 24.5). This phenotype proved to be unstable after passage in mice and sandflies, suggesting that nonfixed alterations are responsible for the elevated resistance. In line with this, single nucleotide polymorphisms and indels identified by whole-genome sequencing could not be directly linked to the decreased drug susceptibility. Copy number variations were absent, whereas somy changes were detected, which may have accounted for the transient acquisition of resistance. Finally, aminopyrazole activity was not influenced by the MDR and MRP efflux pump inhibitors tested. The selection performed does not suggest the rapid development of resistance against aminopyrazoles in the field. Karyotype changes may confer elevated levels of resistance, but these do not seem to be stable in the vertebrate and invertebrate hosts. MDR/MRP efflux pumps are not likely to significantly impact the activity of the aminopyrazole leads.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/AAC.00152-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7449183PMC
August 2020

Interferon Alpha Favors Macrophage Infection by Visceral Species Through Upregulation of Sialoadhesin Expression.

Front Immunol 2020 9;11:1113. Epub 2020 Jun 9.

Laboratory of Microbiology, Parasitology and Hygiene (LMPH), Faculty of Pharmaceutical, Biomedical and Veterinary Sciences, University of Antwerp, Wilrijk, Belgium.

Type I interferons (IFNs) induced by an endogenous RNA virus or exogenous viral infections have been shown to exacerbate infections with New World Cutaneous parasites, however, the impact of type I IFNs in visceral infections and implicated mechanisms remain to be unraveled. This study assessed the impact of type I IFN on macrophage infection with and and the implication of sialoadhesin (Siglec-1/CD169, Sn) as an IFN-inducible surface receptor. Stimulation of bone marrow-derived macrophages with type I IFN (IFN-α) significantly enhanced susceptibility to infection of reference laboratory strains and a set of recent clinical isolates. IFN-α particularly enhanced promastigote uptake. Enhanced macrophage susceptibility was linked to upregulated Sn surface expression as a major contributing factor to the infection exacerbating effect of IFN-α. Stimulation experiments in Sn-deficient macrophages, macrophage pretreatment with a monoclonal anti-Sn antibody or a novel bivalent anti-Sn nanobody and blocking of parasites with soluble Sn restored normal susceptibility levels. Infection of Sn-deficient mice with bioluminescent promastigotes revealed a moderate, strain-dependent role for Sn during visceral infection under the used experimental conditions. These data indicate that IFN-responsive Sn expression can enhance the susceptibility of macrophages to infection with visceral promastigotes and that targeting of Sn may have some protective effects in early infection.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3389/fimmu.2020.01113DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7296180PMC
April 2021

Evaluation of a pan-Leishmania SL RNA qPCR assay for parasite detection in laboratory-reared and field-collected sand flies and reservoir hosts.

Parasit Vectors 2020 Jun 1;13(1):276. Epub 2020 Jun 1.

Laboratory of Microbiology, Parasitology and Hygiene, University of Antwerp, Wilrijk, Belgium.

Background: In eco-epidemiological studies, Leishmania detection in vectors and reservoirs is frequently accomplished by high-throughput and sensitive molecular methods that target minicircle kinetoplast DNA (kDNA). A pan-Leishmania SYBR green quantitative PCR (qPCR) assay which detects the conserved spliced-leader RNA (SL RNA) sequence was developed recently. This study assessed the SL RNA assay performance combined with a crude extraction method for the detection of Leishmania in field-collected and laboratory-reared sand flies and in tissue samples from hyraxes as reservoir hosts.

Methods: Field-collected and laboratory-infected sand fly and hyrax extracts were subjected to three different qPCR approaches to assess the suitability of the SL RNA target for Leishmania detection. Nucleic acids of experimentally infected sand flies were isolated with a crude extraction buffer with ethanol precipitation and a commercial kit and tested for downstream DNA and RNA detection. Promastigotes were isolated from culture and sand fly midguts to assess whether there was difference in SL RNA and kDNA copy numbers. Naive sand flies were spiked with a serial dilution of promastigotes to make a standard curve.

Results: The qPCR targeting SL RNA performed well on infected sand fly samples, despite preservation and extraction under presumed unfavorable conditions for downstream RNA detection. Nucleic acid extraction by a crude extraction buffer combined with a precipitation step was highly compatible with downstream SL RNA and kDNA detection. Copy numbers of kDNA were found to be identical in culture-derived parasites and promastigotes isolated from sand fly midguts. SL RNA levels were slightly lower in sand fly promastigotes (ΔCq 1.7). The theoretical limit of detection and quantification of the SL RNA qPCR respectively reached down to 10 and 10 parasite equivalents. SL RNA detection in stored hyrax samples was less efficient with some false-negative assay results, most likely due to the long-term tissue storage in absence of RNA stabilizing reagents.

Conclusions: This study shows that a crude extraction method in combination with the SL RNA qPCR assay is suitable for the detection and quantification of Leishmania in sand flies. The assay is inexpensive, sensitive and pan-Leishmania specific, and accordingly an excellent assay for high-throughput screening in entomological research.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/s13071-020-04141-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7268266PMC
June 2020

"My Death Will Not [Be] in Vain": Testimonials from Last Gift Rapid Research Autopsy Study Participants Living with HIV at the End of Life.

AIDS Res Hum Retroviruses 2020 12 24;36(12):1071-1082. Epub 2020 Jun 24.

Department of Medicine, University of California San Diego School of Medicine, AntiViral Research Center, University of California San Diego, San Diego, California, USA.

End-of-life (EOL) HIV cure-related research provides a novel approach to studying HIV reservoirs. The Last Gift is a rapid autopsy research study at the University of California San Diego that enrolls terminally ill people living with HIV (PLWHIV) with a desire to contribute to HIV cure-related research. We conducted in-depth baseline and follow-up interviews with Last Gift study participants. We analyzed interview data applying conventional content analysis. Since summer 2017, 13 participants have been enrolled ( = 11 males and 2 females; aged 45-89 years) and 8 participants interviewed. Terminal illnesses included cancers, heart diseases, and neurodegenerative illnesses. Our analysis revealed five key themes: (1) The Last Gift study has tremendous meaning for participants at the end of their life. (2) HIV-specific altruism was a primary motivator to join the Last Gift study, nested within the context of community, scientific advancement, and moral obligation. (3) Participants did not expect physical benefits yet they perceived emotional/psychological, financial, and societal/scientific benefits. (4) There were minimal participant-perceived risks and concerns. (5) Last Gift participants expressed immense gratitude toward study staff. The Last Gift study provides a framework for ethical HIV cure-related research at EOL and highlighted participants' perspectives, motivations, and experiences. Knowing how PLWHIV understand and experience such studies will remain critical to designing ethical, fully informed HIV cure research protocols that are acceptable to PLWHIV.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1089/AID.2020.0020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7703253PMC
December 2020

Perceptions of Next-of-Kin/Loved Ones About Last Gift Rapid Research Autopsy Study Enrolling People with HIV/AIDS at the End of Life: A Qualitative Interview Study.

AIDS Res Hum Retroviruses 2020 12 25;36(12):1033-1046. Epub 2020 Jun 25.

AntiViral Research Center, University of California San Diego, San Diego, California, USA.

A growing number of people living with HIV/AIDS are participating in HIV cure-related research at the end of life (EOL). Due to the novelty of EOL HIV cure-related research, there is a need to understand how their next-of-kin (NOK) perceive such research. We conducted in-depth interviews with NOK of the Last Gift study participants at the University of California, San Diego. The Last Gift study occurs in the context of the EOL and involves a full body donation. NOK completed two interviews: (1) shortly after the participants' enrollment in the study and (2) following death. We applied thematic analysis to analyze qualitative data. NOK included seven individuals (five males and two females), including two spouses, one ex-partner, one sister, a grandmother/grandfather, and a close friend. Thematic analysis revealed five key themes: (1) NOK viewed the Last Gift program in a positive light and had an accurate overall understanding of the study; (2) NOK identified factors that motivated participants to donate their body to science; (3) NOK identified benefits of the Last Gift program for both the donors and themselves; (4) NOK did not perceive any physical risks or decisional regrets of study but wanted to minimize psychosocial impacts and ensure the dignity of participants at all times; and (5) NOK noted elements that remained essential to the successful implementation of EOL HIV cure-related research, such as early involvement and clear communication. Our study uniquely contributes to increased understanding and knowledge of what is important from the point of view of supportive NOK to ensure successful implementation of EOL HIV cure-related research. More research will be needed to understand perspectives of less supportive NOK.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1089/AID.2020.0025DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7703245PMC
December 2020

Impact of clinically acquired miltefosine resistance by Leishmania infantum on mouse and sand fly infection.

Int J Parasitol Drugs Drug Resist 2020 08 1;13:16-21. Epub 2020 May 1.

Laboratory of Microbiology, Parasitology and Hygiene (LMPH), University of Antwerp, Universiteitsplein 1, B-2610, Wilrijk, Belgium. Electronic address:

Objectives: This study evaluated the implications of clinically acquired miltefosine resistance (MIL-R) by assessing virulence in mice and sand flies to reveal the potential of MIL-R strains to circulate.

Methods: Experimental infections with the MIL-R clinical Leishmania infantum isolate MHOM/FR/2005/LEM5159, having a defect in the LiROS3 subunit of the MIL-transporter, and its syngeneic experimentally reconstituted MIL-S counterpart (LEM5159) were performed in BALB/c mice and Lutzomyia longipalpis and Phlebotomus perniciosus sand flies. In mice, the amastigote burdens in liver and spleen were compared microscopically using Giemsa smears and by bioluminescent imaging. During the sand fly infections, the percentage of infected flies, parasite load, colonization of the stomodeal valve and metacyclogenesis were evaluated. The stability of the MIL-R phenotype after sand fly and mouse passage was determined as well.

Results: The fitness of the MIL-R strain differed between the mouse and sand fly infection model. In mice, a clear fitness loss was observed compared to the LiROS3-reconstituted susceptible strain. This defect could be rescued by episomal reconstitution with a wildtype LiROS3 copy. However, this fitness loss was not apparent in the sand fly vector, resulting in metacyclogenesis and efficient colonization of the stomodeal valve. Resistance was stable after passage in both sand fly and mouse.

Conclusion: The natural MIL-R strain is significantly hampered in its ability to multiply and cause a typical visceral infection pattern in BALB/c mice. However, this LiROS3-deficient strain efficiently produced mature infections and metacyclic promastigotes in the sand fly vector highlighting the transmission potential of this particular MIL-R clinical Leishmania strain.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.ijpddr.2020.04.004DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7215113PMC
August 2020

Comparative evaluation of nucleic acid stabilizing reagents for RNA- and DNA-based Leishmania detection in blood as proxy for visceral burdens.

J Microbiol Methods 2020 06 4;173:105935. Epub 2020 May 4.

Laboratory of Microbiology, Parasitology and Hygiene (LMPH), University of Antwerp, Wilrijk, Belgium. Electronic address:

Background: Molecular detection techniques using peripheral blood are preferred over invasive tissue aspiration for the diagnosis and post-treatment follow-up of visceral leishmaniasis (VL) patients. This study aims to identify suitable stabilizing reagents to prevent DNA and RNA degradation during storage and transport to specialized laboratories where molecular diagnosis is performed.

Methodology: The stabilizing capacities of different commercially available reagents were compared using promastigote-spiked human blood and peripheral blood of Syrian golden hamsters subjected to experimental infection, treatment (miltefosine or aminopyrazole DNDi-1044) and immunosuppression. The impact of various storage temperature conditions was tested in combination with an established kinetoplast DNA (kDNA) qPCR and a recently developed spliced leader RNA (SL-RNA) assay for Leishmania detection.

Principal Findings: Irrespective of the blood type and stabilizer used, threshold (cT) values obtained with the SL-RNA qPCR were systematically lower than those obtained with the kDNA assay, confirming the advantage of the SL-RNA assay over the widely used kDNA assay for low-level Leishmania detection. Peripheral blood parasite levels correlated relatively well with hepatic burdens. RNA protect cell reagent provided the most optimal simultaneous DNA and RNA stabilization in both human and hamster blood. However, this stabilizer requires an erythrocyte lysis step, which can be challenging under field conditions. DNA/RNA shield provides a good alternative for downstream kDNA and SL-RNA assays, especially if sample storage capacity at 4 °C can be guaranteed.

Conclusions/significance: The recommended stabilizing reagents are compatible with RNA- and DNA-based Leishmania detection in peripheral blood in the VL hamster model and spiked human blood. Since molecular detection techniques using peripheral blood are less invasive than microscopic assessment of tissue aspirates, the findings of this study may be applied to human VL clinical studies.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.mimet.2020.105935DOI Listing
June 2020

Performing rapid autopsy for the interrogation of HIV reservoirs.

AIDS 2020 06;34(7):1089-1092

Division of Infectious Diseases and Global Public Health, University of California, San Diego, Stein Clinical Research Building.

: Rapid autopsy at the end of life in people with HIV (PWH) permits the preservation of valuable tissue specimens for subsequent study of HIV reservoirs. At our institution, we have developed a cohort of PWH who consent to a rapid autopsy to gather a wide range of fluids and tissues with the goal of advancing HIV cure research. The protocol for successfully performing these autopsies has required careful thought and development over months and years. We have now successfully performed six rapid autopsies and detail here our steps to build the study cohort, train and staff a team of more than a dozen personnel, and process and preserve hundreds of samples from each autopsy.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1097/QAD.0000000000002546DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7780881PMC
June 2020

In Vitro Growth Inhibition Assays of Leishmania spp.

Methods Mol Biol 2020 ;2116:791-800

Laboratory of Microbiology, Parasitology and Hygiene (LMPH), University of Antwerp, Wilrijk-Antwerp, Belgium.

In vitro growth (inhibition) assays have a dual application, either supporting the discovery of novel drugs or as a monitoring tool of drug resistance in patient isolates. From an experimental design point of view, both are quite different with regard to the infecting Leishmania species and strain, the wide variety of permissive host cells (primary cells versus cell lines), drug exposure times, detection methods and endpoint criteria. Recognizing the need for enhanced assay standardization to decrease interlaboratory variation and improve proper interpretation of results, a detailed description is given of the basic fundamental procedures and requirements for routine in vitro growth of Leishmania spp. with specific focus on the intracellular amastigote susceptibility assay. Although the described experimental procedures focus on visceral Leishmania species, the same assay principles may apply for the cutaneous species as well.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/978-1-0716-0294-2_47DOI Listing
February 2021

Phenotypic adaptations of Leishmania donovani to recurrent miltefosine exposure and impact on sand fly infection.

Parasit Vectors 2020 Feb 22;13(1):96. Epub 2020 Feb 22.

Laboratory of Microbiology, Parasitology and Hygiene (LMPH), University of Antwerp, Antwerp, Belgium.

Background: Since the introduction of miltefosine (MIL) as first-line therapy in the kala-azar elimination programme in the Indian subcontinent, treatment failure rates have been increasing. Since parasite infectivity and virulence may become altered upon treatment relapse, this laboratory study assessed the phenotypic effects of repeated in vitro and in vivo MIL exposure.

Methods: Syngeneic Leishmania donovani lines either or not exposed to MIL were compared for drug susceptibility, rate of promastigote multiplication and metacyclogenesis, macrophage infectivity and behaviour in the sand fly vector, Lutzomyia longipalpis.

Results: Promastigotes of both in vitro and in vivo MIL-selected strains displayed a slightly reduced drug susceptibility that was associated with a reduced MIL-accumulation linked to a lower copy number (disomic state) of chromosome 13 harboring the miltefosine transporter (LdMT) gene. In vitro selected promastigotes showed a lower rate of metacyclogenesis whereas the in vivo derived promastigotes displayed a moderately increased growth rate. Repeated MIL exposure did neither influence the parasite load nor metacyclogenesis in the sand fly vector.

Conclusions: Recurrent in vitro and in vivo MIL exposure evokes a number of very subtle phenotypic and genotypic changes which could make promastigotes less susceptible to MIL without attaining full resistance. These changes did not significantly impact on infection in the sand fly vector.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/s13071-020-3972-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7036194PMC
February 2020

Sustainable livestock systems to improve human health, nutrition, and economic status.

Anim Front 2019 Oct 28;9(4):39-50. Epub 2019 Sep 28.

Department for Public Health and Health Professions, University of Florida, Gainesville, FL.

View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1093/af/vfz041DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6951866PMC
October 2019

Transmission potential of paromomycin-resistant Leishmania infantum and Leishmania donovani.

J Antimicrob Chemother 2020 04;75(4):951-957

Laboratory of Microbiology, Parasitology and Hygiene (LMPH), University of Antwerp, Antwerp, Belgium.

Objectives: Former studies demonstrated quick selection of paromomycin resistance for Leishmania infantum and Leishmania donovani accompanied by increased fitness. The present study aimed to interpret these findings in an epidemiological context by comparing infection of WT and experimentally derived paromomycin-resistant strains in the sand fly vector.

Methods: Depending on the Leishmania species, Lutzomyia longipalpis and Phlebotomus perniciosus or Phlebotomus argentipes sand flies were artificially infected with procyclic promastigotes of WT and paromomycin-resistant L. infantum (MHOM/FR/96/LEM3323-cl4) or L. donovani (MHOM/NP/03/BPK275/0-cl18). The infection rate and gut/stomodeal valve colonization were determined to monitor parasite phenotypic behaviour within the vector. The impact of the previously described gain of fitness in the vertebrate host on infectivity for the vector was assessed by feeding L. longipalpis on Syrian golden hamsters heavily infected with either WT or paromomycin-resistant parasites.

Results: WT and paromomycin-resistant Leishmania of both species behaved similarly in terms of infection and parasite location within the studied sand fly species. Blood feeding on infected hamsters did not reveal differences in acquisition of WT and paromomycin-resistant parasites, despite the higher organ burdens observed for the paromomycin-resistant strain. Strains remained resistant after passage in the vector.

Conclusions: Although paromomycin-resistant parasites show an increased parasite fitness in vitro and in laboratory rodents, the intrinsic infection potential of paromomycin-resistant parasites remains unaltered in the sand fly. Of importance is the fact that paromomycin-resistant Leishmania are able to complete development in the natural vectors and produce stomodeal infection with metacyclic forms, which clearly suggests their potential to spread and circulate in nature.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1093/jac/dkz517DOI Listing
April 2020

In-depth comparison of cell-based methodological approaches to determine drug susceptibility of visceral Leishmania isolates.

PLoS Negl Trop Dis 2019 12 2;13(12):e0007885. Epub 2019 Dec 2.

Laboratory of Microbiology, Parasitology and Hygiene (LMPH), University of Antwerp, Antwerp, Belgium.

Monitoring the drug susceptibility of Leishmania isolates still largely relies on standard in vitro cell-based susceptibility assays using (patient-isolated) promastigotes for infection. Although this assay is widely used, no fully standardized/harmonized protocol is yet available hence resulting in the application of a wide variety of host cells (primary cells and cell lines), different drug exposure times, detection methods and endpoint criteria. Advocacy for standardization to decrease inter-laboratory variation and improve interpretation of results has already repeatedly been made, unfortunately still with unsatisfactory progress. As a logical next step, it would be useful to reach at least some agreement on the type of host cell and basic experimental design for routine amastigote susceptibility determination. The present laboratory study using different L. infantum strains as a model for visceral leishmaniasis species compared primary cells (mouse peritoneal exudate (PEC), mouse bone marrow derived macrophages and human peripheral blood monocyte derived macrophages) and commercially available cell lines (THP-1, J774, RAW) for either their susceptibility to infection, their role in supporting intracellular amastigote multiplication and overall feasibility/accessibility of experimental assay protocol. The major findings were that primary cells are better than cell lines in supporting infection and intracellular parasite multiplication, with PECs to be preferred for technical reasons. Cell lines require drug exposure of >96h with THP-1 to be preferred but subject to a variable response to PMA stimulation. The fast dividing J774 and RAW cells out-compete parasite-infected cells precluding proper assay read-out. Some findings could possibly also be applicable to cutaneous Leishmania strains, but this still needs cross-checking. Besides inherent limitations in a clinical setting, susceptibility testing of clinical isolates may remain problematic because of the reliance on patient-derived promastigotes which may exhibit variable degrees of metacyclogenesis and infectivity.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1371/journal.pntd.0007885DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6907865PMC
December 2019

154 compared to 54 mmol per liter of sodium in intravenous maintenance fluid therapy for adult patients undergoing major thoracic surgery (TOPMAST): a single-center randomized controlled double-blind trial.

Intensive Care Med 2019 10 1;45(10):1422-1432. Epub 2019 Oct 1.

Department of Intensive Care Medicine, Antwerp University Hospital, Wilrijkstraat 10, Edegem, 2650, Antwerp, Belgium.

Purpose: To determine the effects of the sodium content of maintenance fluid therapy on cumulative fluid balance and electrolyte disorders.

Methods: We performed a randomized controlled trial of adults undergoing major thoracic surgery, randomly assigned (1:1) to receive maintenance fluids containing 154 mmol/L (Na154) or 54 mmol/L (Na54) of sodium from the start of surgery until their discharge from the ICU, the occurrence of a serious adverse event or the third postoperative day at the latest. Investigators, caregivers and patients were blinded to the treatment. Primary outcome was cumulative fluid balance. Electrolyte disturbances were assessed as secondary endpoints, different adverse events and physiological markers as safety and exploratory endpoints.

Findings: We randomly assigned 70 patients; primary outcome data were available for 33 and 34 patients in the Na54 and Na154 treatment arms, respectively. Estimated cumulative fluid balance at 72 h was 1369 mL (95% CI 601-2137) more positive in the Na154 arm (p < 0.001), despite comparable non-study fluid sources. Hyponatremia < 135 mmol/L was encountered in four patients (11.8%) under Na54 compared to none under Na154 (p = 0.04), but there was no significantly more hyponatremia < 130 mmol/L (1 versus 0; p = 0.31). There was more hyperchloremia > 109 mmol/L under Na154 (24/35 patients, 68.6%) than under Na54 (4/34 patients, 11.8%) (p < 0.001). The treating clinicians discontinued the study due to clinical or radiographic fluid overload in six patients receiving Na154 compared to one patient under Na54 (excess risk 14.2%; 95% CI - 0.2-30.4%, p = 0.05).

Conclusions: In adult surgical patients, sodium-rich maintenance solutions were associated with a more positive cumulative fluid balance and hyperchloremia; hypotonic fluids were associated with mild and asymptomatic hyponatremia.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/s00134-019-05772-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6773673PMC
October 2019

The livestock vaccine supply chain: Why it matters and how it can help eradicate peste des petits Ruminants, based on findings in Karamoja, Uganda.

Vaccine 2019 10 13;37(43):6285-6290. Epub 2019 Sep 13.

Feed the Future Innovation Lab for Livestock Systems,Department of Environmental and Global Health, and the Center for African Studies, University of Florida, Gainesville, FL, USA.

Understanding factors that hinder vaccination, including logistical and social constraints, is critical to finding the most effective approach for the global eradication of peste des petits ruminants (PPR). Vaccination projects should analyze the supply chain and take it into consideration when planning and creating a vaccination strategy. Adequate supply chain management of the PPR vaccine could lead to reduced cost, increased availability, and the construction of a data platform for other livestock vaccines. Integrating the supply chain of PPR vaccine with other veterinary or health commodities could reduce cost, as well as increase uptake. The use of a thermostable vaccine could potentially have a positive impact on the eradication of PPR in remote areas, such as the Karamoja subregion in Uganda, as it did with rinderpest across Sub Saharan Africa. In terms of vaccine delivery, the use of community animal health workers (CAHWs) could be beneficial in certain areas, such as the Karamoja subregion of Uganda, by alleviating supply chain constraints in the last-mile delivery, as well as increasing coverage and uptake. A gendered approach to livestock vaccines should also be considered, as decision-making power regarding livestock vaccination is gendered in many various contexts. The PPR eradication strategy-as well as other livestock vaccination programs-would be more effective and efficient if the supply chain management were considered as a key component in the process and efforts tailored, accordingly.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.vaccine.2019.09.011DOI Listing
October 2019

Impaired development of a miltefosine-resistant Leishmania infantum strain in the sand fly vectors Phlebotomus perniciosus and Lutzomyia longipalpis.

Int J Parasitol Drugs Drug Resist 2019 12 10;11:1-7. Epub 2019 Sep 10.

Laboratory of Microbiology, Parasitology and Hygiene (LMPH), University of Antwerp, Universiteitsplein 1, B-2610, Wilrijk, Belgium. Electronic address:

Objectives: To gain insight into the propagation of miltefosine (MIL) resistance in visceral leishmaniasis, this laboratory study explored development of resistant parasites with a defective miltefosine transporter (MT) in sand flies.

Methods: Infectivity, colonization of stomodeal valve and metacyclogenesis of a MIL-resistant (MIL-R) Leishmania infantum LEM3323 line with a defective MT were assessed in the natural sand fly vectors Phlebotomus perniciosus and Lutzomyia longipalpis. Given our recent description of partial drug dependency of the MT-deficient line, the impact of MIL pre-exposure on sand fly infectivity was explored as well.

Results: A significant reduction in sand fly infection, stomodeal valve colonization and differentiation into metacyclics (determined by a lower flagellum/cell body length ratio) was observed in both vectors for MIL-R as compared to the isogenic parent MIL-susceptible line. Re-introduction of the wildtype MT gene into MIL-R tended to partially rescue the capacity to infect sand flies. Pre-exposure to MIL did not alter infectivity of the MIL-R line.

Conclusion: The MIL resistant L. infantum LEM3323 line is significantly hampered in its development and transmissibility potential in two sand fly vector species. Additional studies are warranted to evaluate whether this applies to other visceral Leishmania parasites with acquired MIL-resistance.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.ijpddr.2019.09.003DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6804374PMC
December 2019

Need for sustainable approaches in antileishmanial drug discovery.

Parasitol Res 2019 Oct 31;118(10):2743-2752. Epub 2019 Aug 31.

Laboratory of Microbiology, Parasitology and Hygiene (LMPH), University of Antwerp, Universiteitsplein 1, 2610, Wilrijk, Antwerp, Belgium.

Leishmaniasis is a neglected parasitic disease for which the current antileishmania therapeutics are hampered by drug toxicity, high cost, need for parenteral administration, increasing treatment failure rates, and emergence of drug resistance. The R&D pipeline had run fairly dry for several years, but fortunately some new drug candidates are now under (pre)clinical development. Identification of novel drugs will nevertheless remain essential to adequately sustain and improve effective disease control in the future. In this review, a package of standard and accessible R&D approaches is discussed with expansion to some alternative strategies focusing on parasite-host and vector-host interactions.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/s00436-019-06443-2DOI Listing
October 2019

Identification of Peste des Petits Ruminants Transmission Hotspots in the Karamoja Subregion of Uganda for Targeting of Eradication Interventions.

Front Vet Sci 2019 5;6:221. Epub 2019 Jul 5.

Cummings School of Veterinary Medicine, Tufts University, Grafton, MA, United States.

This paper describes an assessment of the patterns of virus circulation in the Karamoja subregion of Uganda conducted to identify the communities that maintain the virus and inform the development of a targeted vaccination strategy. Participatory epidemiological methods were used to develop an operational hypothesis for the patterns of PPR in Karamoja that was subsequently validated through outbreak investigation and genomics. The participatory epidemiological assessment included risk mapping with livestock owners, community animal health workers and veterinarians and indicated there were two critical foci of virus transmission on the Uganda-Kenya border. One was located in two adjacent subcounties of Kotido and Kaabong Districts in northern Karamoja and the other in Loroo subcounty of Amudat District in southern Karamoja. Participants reported that these were locations where outbreaks were usually first observed in Karamoja and subsequently spread to other areas. Following the participatory assessment, surveillance activities were implemented across the Karamoja subregion in 2018. Three outbreak were detected, investigated and sampled. Two outbreaks were located in the northern and one on the southern focus of transmission. No Outbreaks were diagnosed in Karamoja outside of these foci during 2018. Genomics indicated different clusters of viruses were associated with the northern and southern foci that were more closely related to other East African isolates than to each other. This indicates these are two separate systems of virus circulation which should be explicitly addressed in eradication as separate cross-border systems that require integrated cross-border interventions.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3389/fvets.2019.00221DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6624733PMC
July 2019

Immunosuppression of Syrian golden hamsters accelerates relapse but not the emergence of resistance in Leishmania infantum following recurrent miltefosine pressure.

Int J Parasitol Drugs Drug Resist 2019 04 11;9:1-7. Epub 2018 Dec 11.

Laboratory of Microbiology, Parasitology and Hygiene (LMPH), University of Antwerp, Antwerp, Belgium. Electronic address:

Although miltefosine (MIL) has only been approved for the treatment of visceral leishmaniasis (VL) in 2002, its application in monotherapy already led to the development of two confirmed MIL-resistant isolates by 2009. Although liposomal amphotericin B is recommended as first-line treatment in Europe, MIL is still occasionally used in HIV co-infected patients. Since their immune system is incapable of controlling the infection, high parasite burdens and post-treatment relapses are common. Linked to the particular pharmacokinetic profile of MIL, successive treatment of recurrent relapses could in principle facilitate the emergence of drug resistance. This study evaluated the effect of immunosuppression (cyclophosphamide 150 mg/kg once weekly) on the development of MIL-resistance in Syrian golden hamsters infected with Leishmania infantum. The hamsters were treated with MIL (20 mg/kg orally for 5 days) whenever clinical signs of infection or relapse were observed. The immunosuppression resulted in a significant depletion of CD4 lymphocytes and MHCII-expressing cells in peripheral blood, and a concomitant increase in tissue parasite burdens and shorter time to relapse, but the strain's susceptibility upon repeated MIL exposure remained unaltered. This study demonstrates that immunosuppression accelerates the occurrence of relapse without expediting MIL resistance development.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.ijpddr.2018.12.001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6296292PMC
April 2019

Miltefosine enhances the fitness of a non-virulent drug-resistant Leishmania infantum strain.

J Antimicrob Chemother 2019 02;74(2):395-406

Laboratory of Microbiology, Parasitology and Hygiene (LMPH), University of Antwerp, Wilrijk, Belgium.

Objectives: Miltefosine is currently the only oral drug for visceral leishmaniasis, and although deficiency in an aminophospholipid/miltefosine transporter (MT) is sufficient to elicit drug resistance, very few naturally miltefosine-resistant (MIL-R) strains have yet been isolated. This study aimed to make a detailed analysis of the impact of acquired miltefosine resistance and miltefosine treatment on in vivo infection.

Methods: Bioluminescent versions of a MIL-R strain and its syngeneic parental line were generated by integration of the red-shifted firefly luciferase PpyRE9. The fitness of both lines was compared in vitro (growth rate, metacyclogenesis and macrophage infectivity) and in BALB/c mice through non-invasive bioluminescence imaging under conditions with and without drug pressure.

Results: This study demonstrated a severe fitness loss of MT-deficient parasites, resulting in a complete inability to multiply and cause a typical visceral leishmaniasis infection pattern in BALB/c mice. The observed fitness loss could not be rescued by host immune suppression with cyclophosphamide, whereas episomal reconstitution with a wild-type MT restored parasite virulence, hence linking parasite fitness to MT mutation. Remarkably, in vivo miltefosine treatment or in vitro miltefosine pre-exposure significantly rescued MIL-R parasite virulence. The in vitro pre-exposed MIL-R promastigotes showed a longer and more slender morphology, suggesting an altered membrane composition.

Conclusions: The profound fitness loss of MT-deficient parasites most likely explains the low frequency of MIL-R clinical isolates. The observation that miltefosine can reverse this phenotype indicates a drug dependency of the MT-deficient parasites and emphasizes the importance of resistance profiling prior to miltefosine administration.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1093/jac/dky450DOI Listing
February 2019

Impact of primary mouse macrophage cell types on Leishmania infection and in vitro drug susceptibility.

Parasitol Res 2018 Nov 23;117(11):3601-3612. Epub 2018 Aug 23.

Faculty of Pharmaceutical, Biomedical and Veterinary Sciences, Laboratory of Microbiology, Parasitology and Hygiene, University of Antwerp, Wilrijk, Belgium.

Primary mouse macrophages are frequently used to provide an in vitro intracellular model to evaluate antileishmanial drug efficacy. The present study compared the phenotypic characteristics of Swiss, BALB/c, and C57BL/6 mouse bone marrow-derived macrophages and peritoneal exudate cells using different stimulation and adherence protocols upon infection with a Leishmania infantum laboratory strain and two clinical isolates. Evaluation parameters were susceptibility to infection, permissiveness to amastigote multiplication, and impact on drug efficacy. Observed variations in infection of peritoneal exudate cells can mostly be linked to changes in the inflammatory cytokine profiles (IL-6, TNF-α, KC/GRO) rather than to differences in initial production of nitric oxide and reactive oxygen species. Optimization of the cell stimulation and adherence conditions resulted in comparable infection indices among peritoneal exudate cells and the various types of bone marrow-derived macrophages. BALB/c-derived bone marrow-derived macrophages were slightly more permissive to intracellular amastigote replication. Evaluation of antileishmanial drug potency in the various cell systems revealed minimal variation for antimonials and paromomycin, and no differences for miltefosine and amphotericin B. The study results allow to conclude that drug evaluation can be performed in all tested primary macrophages as only marginal differences are observed in terms of susceptibility to infection and impact of drug exposure. Combined with some practical considerations, the use of 24-h starch-stimulated, 48-h adhered, Swiss-derived peritoneal exudate cells can be advocated as an efficient, reliable, relatively quick, and cost-effective tool for routine drug susceptibility testing in vitro whenever the use of primary cells is feasible.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/s00436-018-6059-4DOI Listing
November 2018

App-based learning as an alternative for instructors in teaching basic life support to school children: a randomized control trial.

Acta Clin Belg 2019 Oct 14;74(5):317-325. Epub 2018 Aug 14.

Department of Emergency Medicine, University Hospitals Leuven , Leuven , Belgium.

: The importance of early cardiopulmonary resuscitation (CPR) during an out-of-hospital cardiac arrest (OHCA) cannot be emphasized enough and has a major impact on survival. Unfortunately, CPR education in schools is often inadequate or non-existing due to lack of educators and financial resources. The introduction of application-based teaching could facilitate education as no instructor is needed. The aim of present study is to compare app-based self-teaching (intervention group) with a traditional instructor-led course (control group). This trial is in line with the endorsement of training school children by the World Health Organization (WHO). : This randomized controlled trial contains 165 participants between 16 and 18 years. The control group followed an instructor-led course, while the intervention group used a tablet application to complete a self-education course.Both groups were taught in separated classes during 40 min and used manikins and automated external defibrillator (AED) trainer kits. Before and after training, both groups were evaluated in a practical test to examine their skills (an overall score and sub-scores on different sections were obtained, following ERC 2015 guidelines). : No significant difference ( = 0.304) in overall teaching effectiveness was found between both groups. Although, in a sub-analysis, we found significantly better results for the instructor group for checking airway ( = 0.018), asking for an AED ( < 0.01) and shocking the patient ( = 0.002). : App-based teaching can be a valuable alternative for classic instructor-based education. Yet new apps have to be evaluated before being implemented in CPR education.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1080/17843286.2018.1500766DOI Listing
October 2019

Cyclic Nucleotide-Specific Phosphodiesterases as Potential Drug Targets for Anti-Leishmania Therapy.

Antimicrob Agents Chemother 2018 10 24;62(10). Epub 2018 Sep 24.

Centro de Investigaciones Biológicas (CIB, CSIC), Madrid, Spain

The available treatments for leishmaniasis are less than optimal due to inadequate efficacy, toxic side effects, and the emergence of resistant strains, clearly endorsing the urgent need for discovery and development of novel drug candidates. Ideally, these should act via an alternative mechanism of action to avoid cross-resistance with the current drugs. As cyclic nucleotide-specific phosphodiesterases (PDEs) of have been postulated as putative drug targets, a series of potential inhibitors of PDEs were explored. Several displayed potent and selective activity against intracellular amastigotes. One imidazole derivative, compound 35, was shown to reduce the parasite loads and to increase the cellular cyclic AMP (cAMP) level at in a dose-dependent manner at just 2× and 5× the 50% inhibitory concentration (IC), indicating a correlation between antileishmanial activity and increased cellular cAMP levels. Docking studies and molecular dynamics simulations pointed to imidazole 35 exerting its activity through PDE inhibition. This study establishes for the first time that inhibition of cAMP PDEs can potentially be exploited for new antileishmanial chemotherapy.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/AAC.00603-18DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6153811PMC
October 2018

In vitro and in vivo pharmacodynamics of three novel antileishmanial lead series.

Int J Parasitol Drugs Drug Resist 2018 04 31;8(1):81-86. Epub 2018 Jan 31.

Laboratory for Microbiology, Parasitology and Hygiene (LMPH), University of Antwerp, Belgium. Electronic address:

Objectives: Three new chemical series (bicyclic nitroimidazoles, aminopyrazoles and oxaboroles) were selected by Drugs for Neglected Diseases initiative as potential new drug leads for leishmaniasis. Pharmacodynamics studies included both in vitro and in vivo efficacy, cross-resistance profiling against the current antileishmanial reference drugs and evaluation of their cidal activity potential.

Methods: Efficacy against the reference laboratory strains of Leishmania infantum (MHOM/MA(BE)/67/ITMAP263) and L. donovani (MHOM/ET/67/L82) was evaluated in vitro on intracellular amastigotes and in vivo in the early curative hamster model. Cidal activity was assessed over a period of 15 days in an in vitro 'time-to-kill' assay. Cross-resistance was assessed in vitro on a panel of L. infantum strains with different degrees of resistance to either antimony, miltefosine or paromomycin.

Results: All lead compounds showed potent and selective in vitro activity against the Leishmania strains tested and no cross-resistance could be demonstrated against any of the current antileishmanial drugs. Cidal activity was obtained in vitro for all series within 15 days of exposure with some differences noted between L. donovani and L. infantum. When evaluated in vivo, all lead compounds showed high efficacy and no adverse effects were observed.

Conclusions: The new lead series were shown to have cidal pharmacodynamic activity. The absence of cross-resistance with any of the current antileishmanial drugs opens possibilities for combination treatment to reduce the likelihood of treatment failures and drug resistance.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.ijpddr.2018.01.006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6114106PMC
April 2018

Report of antileishmanial properties of Iberian macroalgae.

Nat Prod Res 2019 Jun 9;33(12):1778-1782. Epub 2018 Feb 9.

a Centre of Marine Sciences , University of Algarve , Faro , Portugal.

Here is reported the anti activity of 48 hexane, CHCl and MeOH extracts from 16 macroalgae collected on the Iberian Coast. Seven hexane and CHCl and extracts were active towards promastigotes (IC 29.8-101.8 μg/mL) inducing strong morphological alterations in the parasites. Hexane extracts of and were also active against intracellular amastigotes (IC 5.1 and 6.8 μg/mL, respectively). Fatty acids, triacylglycerols, carotenoids, steroids and meroterpenoids were detected by nuclear magnetic resonance (NMR), and gas chromatography in the extracts. These results suggest that macroalgae contain compounds with antileishmanial activity, which could be explored as scaffolds to the development of novel sources of antiparasitic derivatives.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1080/14786419.2018.1434637DOI Listing
June 2019

Elongator subunit 3 (ELP3) modifies ALS through tRNA modification.

Hum Mol Genet 2018 04;27(7):1276-1289

Department of Neurosciences, Experimental Neurology and Leuven Institute for Neuroscience and Disease (LIND), KU Leuven-University of Leuven, B-3000 Leuven, Belgium.

Amyotrophic lateral sclerosis (ALS) is a fatal degenerative motor neuron disorder of which the progression is influenced by several disease-modifying factors. Here, we investigated ELP3, a subunit of the elongator complex that modifies tRNA wobble uridines, as one of such ALS disease modifiers. ELP3 attenuated the axonopathy of a mutant SOD1, as well as of a mutant C9orf72 ALS zebrafish model. Furthermore, the expression of ELP3 in the SOD1G93A mouse extended the survival and attenuated the denervation in this model. Depletion of ELP3 in vitro reduced the modified tRNA wobble uridine mcm5s2U and increased abundance of insoluble mutant SOD1, which was reverted by exogenous ELP3 expression. Interestingly, the expression of ELP3 in the motor cortex of ALS patients was reduced and correlated with mcm5s2U levels. Our results demonstrate that ELP3 is a modifier of ALS and suggest a link between tRNA modification and neurodegeneration.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1093/hmg/ddy043DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6159532PMC
April 2018

Evaluation of a Pan-Leishmania Spliced-Leader RNA Detection Method in Human Blood and Experimentally Infected Syrian Golden Hamsters.

J Mol Diagn 2018 03 31;20(2):253-263. Epub 2018 Jan 31.

Laboratory of Microbiology, Parasitology and Hygiene, University of Antwerp, Wilrijk, Belgium. Electronic address:

Several methods have been developed for the detection of Leishmania, mostly targeting the minicircle kinetoplast DNA (kDNA). A new RNA real-time quantitative PCR (qPCR) assay was developed targeting the conserved and highly expressed spliced-leader (SL) mini-exon sequence. This study compared the limits of detection of various real-time PCR assays in hamsters infected with Leishmania infantum, in spiked human blood, and in clinical blood samples from visceral leishmaniasis patients. The SL-RNA assay showed an excellent analytical sensitivity in tissues (0.005 and 0.002 parasites per mg liver and spleen, respectively) and was not prone to false-positive reactions. Evaluation of the SL-RNA assay on clinical samples demonstrated lower threshold cycle values than the kDNA qPCR, an excellent interrun stability of 97%, a 93% agreement with the kDNA assay, and an estimated sensitivity, specificity, and accuracy of 93.2%, 94.3%, and 93.8%, respectively. The SL-RNA qPCR assay was equally efficient for detecting Leishmania major, Leishmania tropica, Leishmania mexicana, Leishmania guayensis, Leishmania panamensis, Leishmania braziliensis, L. infantum, and Leishmania donovani and revealed similar SL-RNA levels in the different species and the occurrence of polycistronic SL-containing transcripts in Viannia species. Collectively, this single SL-RNA qPCR assay enables universal Leishmania detection and represents a particularly useful addition to the widely used kDNA assay in clinical studies in which the detection of viable parasites is pivotal to assess parasitological cure.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.jmoldx.2017.12.003DOI Listing
March 2018