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Author: Donna Huber

Survival and Internalization of Salmonella and Escherichia coli O157:H7 Sprayed onto Different Cabbage Cultivars during Cultivation in Growth Chambers

Donna Huber on January 9, 2019

cabbage

ABSTRACT

BACKGROUND: Cabbage may become contaminated with enteric pathogens during cultivation. Using multiple cabbage cultivars at two maturity stages (small plants or plants with small heads) in growth chamber studies, the fate (internalization or surface survival) of Salmonella and Escherichia coli O157:H7 (0157) were examined in conjunction with any potential relationships to the plant’s antimicrobial content.

RESULTS: Internalized Salmonella was detected in cabbage within 24 hours with prevalence ranging from 62% (16 of 26) for the ‘Super Red 80’ cultivar to 92% (24 of 26) for the ‘Red Dynasty’ cultivar. The fate of Salmonella and O157 on small cabbage plants over nine days was significantly affected by cultivar with both these pathogens surviving the least and most on the ‘Capture’ and ‘Farao’ cultivars, respectively (P < 0.05). Survival of O157 was slightly higher on cabbage heads for O157 than small plants suggesting that the maturity stage may affect this pathogen’s fate. An inverse relationship existed between antimicrobial levels and a pathogen’s fate on cabbage heads (P < 0.05).

CONCLUSIONS: The fate of pathogens varied with the cabbage cultivar in growth chamber studies highlighting the potential to explore cultivar in field studies to reduce the risk of microbiological contamination in this crop.

Marilyn C. Erickson, Jye-Yin Liao, Alison S. Payton, Peter W. Cook, Ynes R. Ortega. 2019. Journal of the Science of Food and Agriculture.
https://doi.org/10.1002/jsfa.9573

Researchers receive $2M NIH instrumentation grant

Donna Huber on January 8, 2019

by Alan Flurry

The National Institutes of Health has awarded University of Georgia researchers $1.956 million for a high-resolution mass spectrometer that will enhance capabilities for scientists in many fields across campus.

The award by the NIH High End Instrumentation program, which provides grants in the range of $600,000 to $2 million for a variety of expensive instrumentation, including MRI imagers, electron microscopes, DNA sequencers, and mass spectrometers, was one of 30 awards made in the program, and one of only six mass spectrometer requests funded in the 2018 cycle.

The grant funded a 12 Tesla Bruker Solarix FTMS, a high-resolution mass spectrometer capable of measuring molecular weights with precision accuracy that can be applied to molecules ranging in size from small metabolic products to intact proteins and protein complexes. It can also provide molecular structure through a multidimensional analysis method known as tandem mass spectrometry. The instrument will be used to support research in metabolomics and glycomics, the analysis of genetic, physiologic and pathologic aspects of sugar molecules involved in all biological process from modulating cell function to determining cancer development.

“This instrument will enhance the research capabilities for a number of scientists in chemistry, the biological sciences and biomedical research, and will help foster interdisciplinary research projects between groups in a number of departments and colleges at the university,” said Jon Amster, professor and head of the department of chemistry and principal investigator on the grant.

Over a dozen researchers will be major users of this instrument, which will be housed in the Amster laboratory in the department of chemistry.

“The new 12T FT-ICR instrument will greatly improve our ability to perform metabolomics analysis, especially regarding to the identification of unknown metabolites, since this instrument has higher accuracy and resolving power than the current instruments at UGA,” said Belen Cassera, associate professor of biochemistry and molecular biology, member of the Center for Tropical and Emerging Global Diseases, and co-principal investigator on the grant. “This type of grant can be particularly difficult to obtain and it is a privilege for me to be part of an amazing team of investigators that put together this application.”

“Virtually every metabolomics project we have going right now will benefit from this new instrumentation grant,” said Art Edison, GRA Eminent Scholar, professor of biochemistry and molecular biology, and a co-principal investigator on the grant. “High resolution mass spectrometry is a very important tool for the analysis of complex biological mixtures and unknown metabolite identification in applications ranging from human disease to carbon cycling in the ocean to model organisms for pathway analysis.”

Of the 104 NIH shared instrumentation grants made this year during 2018, only 10 were in the range of $1.9 million to $2 million.

Thirty-Day Daily Comparisons of Kato-Katz and CCA Assays of 45 Egyptian Children in Areas with Very Low Prevalence of Schistosoma mansoni

Donna Huber on January 2, 2019

Egyptian children

Forty-five Schistosoma mansoni egg–negative/circulating cathodic antigen (CCA) low (Trace-1+) positive children in areas of very low prevalence were followed up daily for 30 days. Stool and urine specimens were collected and examined each day from each child. At the midpoint of the study, three egg-positive control persons with light intensity infection were included in the protocol. Stool samples were examined by the Kato–Katz (four slides/stool sample) technique and all S. mansoni egg–negative stools were further tested by the “miracidia hatching test” (MHT). Urine samples were examined by the point-of-care CCA assay (POC-CCA). Over 30 days, only one of 1,338 consecutive stool samples from study subjects was S. mansoni egg and MHT positive (0.07%). Egg counts fluctuated daily in stools from positive controls and S. mansoni miracidia were detected in all but two samples by the MHT. Point-of-care–circulating cathodic antigen bands were scored from G1 to G10 and then translated to standard Trace, 1+, 2+, 3+ banding patterns. In two districts, the POC-CCA assays were Trace or 1+ for both the study children and the positive controls. In the third district, the POC-CCA assays were Trace or 1+ for the study children and 1+ or 2+ for the positive control. We conclude that in areas with extremely low prevalence S. mansoni egg–negative and CCA-Trace or 1+ children are unlikely to pose substantial risks to continued transmission of schistosomiasis. In this setting, POC-CCA Trace or 1+ readings are likely to be false positives or perhaps represent low-level single-sex schistosome infections.

Ayat A. Haggag, Amal Rabiee, Khaled M. Abd Elaziz, Carl H. Campbell Jr., Daniel G. Colley, and Reda M. R. Ramzy. 2019 The American Journal of Tropical Medicine and Hygiene. https://doi.org/10.4269/ajtmh.18-0829

Efficacy and side effects of doxycycline versus minocycline in the three-dose melarsomine canine adulticidal heartworm treatment protocol

Donna Huber on December 27, 2018

Abstract

Background: The American Heartworm Society currently recommends the use of a monthly macrocyclic lactone, a 28-day course of 10 mg/kg doxycycline BID, and the 3-dose protocol of melarsomine dihydrochloride for the treatment of canine heartworm disease. Doxycycline is necessary for the reduction of the bacterium Wolbachia, found in all heartworm life-stages. Previous price increases and decreasing availability prompted us to evaluate alternative tetracycline antibiotics, i.e. minocycline, for the reduction of Wolbachia during canine heartworm treatment.

Methods: Thirty-two heartworm-positive dogs were randomized to receive 10 mg/kg or 5 mg/kg of either doxycycline or minocycline for 28 days BID, for a total of 8 dogs per experimental group. All dogs received 6 months of Heartgard Plus® (ivermectin/pyrantel) and the 3-dose protocol of 2.5 mg/kg melarsomine dihydrochloride. Blood samples were collected prior to the initiation of treatment, every 7 days throughout tetracycline treatment, and then monthly thereafter until the dog tested negative for the presence of heartworm antigen. DNA was isolated from circulating microfilarial samples and qPCR was performed on each sample.

Results: A greater number of dogs in the 10 mg/kg doxycycline and minocycline treated groups experienced gastrointestinal side effects as compared to the 5 mg/kg doxycycline and minocycline treated groups. All eight dogs in the 10 mg/kg doxycycline-treated group tested negative for the presence of Wolbachia DNA by 28 days post-tetracycline treatment. A total of two dogs in both the 5 mg/kg doxycycline- and 10 mg/kg minocycline-treated groups and three dogs in the 5 mg/kg minocycline-treated group remained positive for the presence of Wolbachia DNA by the end of tetracycline treatment.

Conclusions: No lung pathology was assessed in this clinical trial, therefore the clinical effect of the remaining Wolbachia DNA in the 10 mg/kg minocycline-, 5 mg/kg doxycycline- and 5 mg/kg minocycline-treated groups cannot be determined. Owner compliance in the proper administration of these tetracyclines may be impacted by the increased severe gastrointestinal side effects reported for the 10 mg/kg doxycycline- and minocycline-treated groups. We recommend that veterinarians prescribe the recommended 10 mg/kg doxycycline for canine heartworm treatment and reduce the dosage to 5 mg/kg in cases of severe gastrointestinal side effects in order to improve owner compliance in administration of medications.

Molly D. Savadelis, Katherine M. Day, Jenna L. Bradner, Adrian J. Wolstenholme, Michael T. Dzimianski, and Andrew R. Moorhead. 2018. Parasites & Vectors; 11:671. https://doi.org/10.1186/s13071-018-3264-z

Translating preventive chemotherapy prevalence thresholds for Schistosoma mansoni from the Kato-Katz technique into the point-of-care circulating cathodic antigen diagnostic test

Donna Huber on December 14, 2018

The World Health Organization (WHO) has defined goals for schistosomiasis morbidity control to be reached by 2025 that are based on preventive chemotherapy. Intervention thresholds for Schistosoma mansoni are currently defined for prevalence measured by stool microscopy using the Kato-Katz technique. However, the Kato-Katz technique shows low sensitivity, particularly for the detection of light-intensity infections. Replacing it with the semi-quantitative point-of-care circulating cathodic antigen (POC-CCA) urine cassette test requires translation of the thresholds and precise characterization of the diagnostic sensitivity and specificity. In this study, we applied a novel egg-count model to a suite of data obtained from different settings in Africa and the Americas with diverse endemicity levels. We used a simulation study to infer on the relation between Kato-Katz and POC-CCA prevalence. Based on our study, we were able to provide recommendations for POC-CCA thresholds taking into account semi-quantitative results of the test. We found that a S. mansoni prevalence of 10% based on duplicate slide Kato-Katz thick smear is equivalent to 15–40% POC-CCA prevalence when trace results are considered positive and to 10–20% POC-CCA prevalence when trace results are considered negative. Our results have important bearings for mapping, control, surveillance, and verification of elimination of intestinal schistosomiasis.

Oliver Bärenbold, Amadou Garba, Daniel G. Colley, Fiona M. Fleming, Ayat A. Haggag, Reda M. R. Ramzy, Rufin K. Assaré, Edridah M. Tukahebwa, Jean B. Mbonigaba, Victor Bucumi, Biruck Kebede, Makoy S. Yibi, Aboulaye Meité, Jean T. Coulibaly, Eliézer K. N’Goran, Louis-Albert Tchuem Tchuenté, Pauline Mwinzi, Jürg Utzinger, Penelope Vounatsou. 2018. PLOS Neglected Tropical Diseases. https://doi.org/10.1371/journal.pntd.0006941

Drug Discovery for Kinetoplastid Diseases: Future Directions

Donna Huber on December 13, 2018

Kinetoplastid parasites have caused human disease for millennia. Significant achievements have been made toward developing new treatments for leishmaniasis (particularly on the Indian subcontinent) and for human African trypanosomiasis (HAT). Moreover, the sustained decrease in the incidence of HAT has made the prospect of elimination a tantalizing reality. Despite the gains, no new chemical or biological entities to treat kinetoplastid diseases have been registered in more than three decades, and more work is needed to discover safe and effective therapies for patients with Chagas disease and leishmaniasis. Advances in tools for drug discovery and novel insights into the biology of the host–parasite interaction may provide opportunities for accelerated progress. Here, we summarize the output from a gathering of scientists and physicians who met to discuss the current status and future directions in drug discovery for kinetoplastid diseases.

Srinivasa P. S. Rao, Michael P. Barrett, Glenn Dranoff, Christopher J. Faraday, Claudio R. Gimpelewicz, Asrat Hailu, Catherine L. Jones, John M. Kelly, Janis K. Lazdins-Helds, Pascal Mäser, Jose Mengel, Jeremy C. Mottram, Charles E. Mowbray, David L. Sacks, Phillip Scott &, Gerald F. Späth, Rick L. Tarleton, Jonathan M. Spector, and Thierry T. Diagana. 2018. ACS Infectious Diseases. https://pubs.acs.org/doi/10.1021/acsinfecdis.8b00298

Evaluation of a Single Dose of Azithromycin for Trachoma in Low-Prevalence Communities

Donna Huber on December 13, 2018

ABSTRACT

Purpose: Trachoma, caused by repeated ocular infection with Chlamydia trachomatis, is the leading infectious cause of blindness worldwide and is targeted for elimination as a public health problem. We sought to determine whether a one-time azithromycin mass treatment would reduce trachomatous inflammation–follicular (TF) levels below the elimination threshold of 5% in communities with disease prevalence between 5 and 9.9%.

Methods: The study was conducted in 96 sub-village units (balozis) in the Kongwa district of Tanzania which were predicted from prior prevalence surveys to have TF between 5 and 9.9%. Balozis were randomly assigned to the intervention and control arms. The intervention arm received a single mass drug administration of azithromycin. At baseline and 12-month follow-up, ocular exams for trachoma, ocular swabs for detection of chlamydial DNA, and finger prick blood for analysis of anti-chlamydial antibody were taken.

Results: Comparison of baseline and 12-month follow-up showed no significant difference in the overall TF1-9 prevalence by balozi between control and treatment arms. In the treatment arm there was a significant reduction of ocular infection 12 months after treatment (p = 0.004) but no change in the control arm. No change in Pgp3-specific antibody responses were observed after treatment in the control or treatment arms. Anti-CT694 responses increased in both study arms (p = 0.009 for control arm and p = 0.04 for treatment arm).

Conclusion: These data suggest that a single round of MDA may not be sufficient to decrease TF levels below 5% when TF1-9 is between 5 and 9.9% at baseline.

Nana Wilson, Brook Goodhew, Harran Mkocha, Kahaliah Joseph, Claudiu Bandea, Carolyn Black, Joseph Igietseme, Beatriz Munoz, Sheila K. West, Patrick Lammie, Mabula Kasubi & Diana L. Martin. 2019. Ophthalmic Epidemiology; 26(1):1-6. DOI: 10.1080/09286586.2017.1293693

Open-source discovery of chemical leads for next-generation chemoprotective antimalarials

Donna Huber on December 7, 2018

Abstract

To discover leads for next-generation chemoprotective antimalarial drugs, we tested more than 500,000 compounds for their ability to inhibit liver-stage development of luciferase-expressing Plasmodium spp. parasites (681 compounds showed a half-maximal inhibitory concentration of less than 1 micromolar). Cluster analysis identified potent and previously unreported scaffold families as well as other series previously associated with chemoprophylaxis. Further testing through multiple phenotypic assays that predict stage-specific and multispecies antimalarial activity distinguished compound classes that are likely to provide symptomatic relief by reducing asexual blood-stage parasitemia from those which are likely to only prevent malaria. Target identification by using functional assays, in vitro evolution, or metabolic profiling revealed 58 mitochondrial inhibitors but also many chemotypes possibly with previously unidentified mechanisms of action.

Yevgeniya Antonova-Koch, Stephan Meister, Matthew Abraham, Madeline R. Luth, Sabine Ottilie, Amanda K. Lukens, Tomoyo Sakata-Kato, Manu Vanaerschot, Edward Owen, Juan Carlos Jado, Steven P. Maher, Jaeson Calla, David Plouffe, Yang Zhong, Kaisheng Chen, Victor Chaumeau, Amy J. Conway, Case W. McNamara, Maureen Ibanez, Kerstin Gagaring, Fernando Neria Serrano, Korina Eribez, Cullin McLean Taggard, Andrea L. Cheung, Christie Lincoln, Biniam Ambachew, Melanie Rouillier, Dionicio Siegel, François Nosten, Dennis E. Kyle, Francisco-Javier Gamo, Yingyao Zhou, Manuel Llinás, David A. Fidock, Dyann F. Wirth, Jeremy Burrows, Brice Campo, Elizabeth A. Winzeler. 2018. Science; 362(6419):eaat9446. http://science.sciencemag.org/content/362/6419/eaat9446

Cryogenically preserved RBCs support gametocytogenesis of Plasmodium falciparum in vitro and gametogenesis in mosquitoes

Donna Huber on December 6, 2018

Abstract

Background: The malaria Eradication Research Agenda (malERA) has identified human-to-mosquito transmission of Plasmodium falciparum as a major target for eradication. The cornerstone for identifying and evaluating transmission in the laboratory is standard membrane feeding assays (SMFAs) where mature gametocytes of P. falciparum generated in vitro are offered to mosquitoes as part of a blood-meal. However, propagation of “infectious” gametocytes requires 10–12 days with considerable physico-chemical demands imposed on host RBCs and thus, “fresh” RBCs that are ≤ 1-week old post-collection are generally recommended. However, in addition to the costs, physico-chemical characteristics unique to RBC donors may confound reproducibility and interpretation of SMFAs. Cryogenic storage of RBCs (“cryo-preserved RBCs”) is accepted by European and US FDAs as an alternative to refrigeration (4 °C) for preserving RBC “quality” and while cryo-preserved RBCs have been used for in vitro cultures of other Plasmodia and the asexual stages of P. falciparum, none of the studies required RBCs to support parasite development for > 4 days.

Results: Using the standard laboratory strain, P. falciparum NF54, 11 SMFAs were performed with RBCs from four separate donors to demonstrate that RBCs cryo-preserved in the gaseous phase of liquid nitrogen (− 196 °C) supported gametocytogenesis in vitro and subsequent gametogenesis in Anopheles stephensimosquitoes. Overall levels of sporogony in the mosquito, as measured by oocyst and sporozoite prevalence, as well as oocyst burden, from each of the four donors thawed after varying intervals of cryopreservation (1, 4, 8, and 12 weeks) were comparable to using ≤ 1-week old refrigerated RBCs. Lastly, the potential for cryo-preserved RBCs to serve as a suitable alternative substrate is demonstrated for a Cambodian isolate of P. falciparum across two independent SMFAs.

Conclusions: Basic guidelines are presented for integrating cryo-preserved RBCs into an existing laboratory/insectary framework for P. falciparum SMFAs with significant potential for reducing running costs while achieving greater reliability. Lastly, scenarios are discussed where cryo-preserved RBCs may be especially useful in enhancing the understanding and/or providing novel insights into the patterns and processes underlying human-to-mosquito transmission.

Ashutosh K. Pathak, Justine C. Shiau, Matthew B. Thomas and Courtney C. Murdock. 2018. Malaria Journal; 17:457. https://doi.org/10.1186/s12936-018-2612-y

Proximal Remote Sensing to Non-destructively Detect and Diagnose Physiological Responses by Host Insect Larvae to Parasitism

Donna Huber on December 4, 2018

As part of identifying and characterizing physiological responses and adaptations by insects, it is paramount to develop non-destructive techniques to monitor individual insects over time. Such techniques can be used to optimize the timing of when in-depth (i.e., destructive sampling of insect tissue) physiological or molecular analyses should be deployed. In this article, we present evidence that hyperspectral proximal remote sensing can be used effectively in studies of host responses to parasitism. We present time series body reflectance data acquired from individual soybean loopers (Chrysodeixis includens) without parasitism (control) or parasitized by one of two species of parasitic wasps with markedly different life histories: Microplitis demolitor, a solitary larval koinobiont endoparasitoid and Copidosoma floridanum, a polyembryonic (gregarious) egg-larval koinobiont endoparasitoid. Despite considerable temporal variation in reflectance data 1–9 days post-parasitism, the two parasitoids caused uniquely different host body reflectance responses. Based on reflectance data acquired 3–5 days post-parasitism, all three treatments (control larvae, and those parasitized by either M. demolitor or C. floridanum) could be classified with >85 accuracy. We suggest that hyperspectral proximal imaging technologies represent an important frontier in insect physiology, as they are non-invasive and can be used to account for important time scale factors, such as: minutes of exposure or acclimation to abiotic factors, circadian rhythms, and seasonal effects. Although this study is based on data from a host-parasitoid system, results may be of broad relevance to insect physiologists. Described approaches provide a non-invasive and rapid method that can provide insights into when to destructively sample tissue for more detailed mechanistic studies of physiological responses to stressors and environmental conditions.

Christian Nansen and Michael R. Strand. 2018. Frontiers in Physiology. https://doi.org/10.3389/fphys.2018.01716