Portable PCR field-based detection of sweetpotato viruses

Sweetpotato ( Ipomoea batatas Lam.) production is greatly constrained by viral infections, especially Sweet potato feathery mottle virus and Sweet potato chlorotic stunt virus that synergistically cause a severe sweetpotato virus disease. The impact of viruses is aggravated by the vegetative nature of the crop and inaccessibility to dependable diagnostic tools in rural areas where sweetpotato production is done. This makes it hard for seed inspectors to perform quality checks prior to use of vines for planting. The objective of this study was to develop a procedure that allows for detection of sweetpotato viruses on-site. This involved modification of the Lodhi et al. (1994) nucleic acid extraction procedure, by omitting some of the laboratory specific steps and varying the incubation time in liquid nitrogen, instead of the freezer. Incubation in liquid nitrogen for only 1.5 hours yielded as high quality RNA compared to that of the original protocol, when incubation was done at 4\ub0C overnight in a freezer. Reverse transcriptase (RT) was run using a portable miniPCR thermocycler; and the resulting cDNA was amplified using this miniPCR machine instead of using a laboratory stationed conventional PCR thermocycler. The cDNA was efficiently amplified and amplicons were similar to those obtained with the original extraction protocol and subsequent amplification by the conventional RT-PCR. Our protocol reduced extraction time from about 16 hours for the original protocol, to about 2 hours and 45 minutes. If this tool is utilised by the crop protection departments, we believe it will contribute greatly towards sustainable sweetpotato production through making timely recommendations.La production de la patate douce ( Ipomoea batatas Lam.) est fortement limit\ue9e par les infections virales, en particulier le virus de la marbrure plumeuse de la patate douce et le virus du stunt chlorotique de la patate douce qui provoquent en synergie une maladie virale grave de la patate douce. L\u2019impact des virus est aggrav\ue9 par la nature v\ue9g\ue9tative de la culture et l\u2019inaccessibilit\ue9 des outils fiables pour le diagnostic dans les zones rurales o\uf9 la production de patate douce est r\ue9alis\ue9e. Cela rend difficile les inspecteurs des semences d\u2019effectuer des contr\uf4les de qualit\ue9 avant l\u2019utilisation des vignes par les agriculteurs. L\u2019objectif de cette \ue9tude \ue9tait de d\ue9velopper une proc\ue9dure permettant la d\ue9tection des virus de la patate douce sur place. Cela impliquait une modification de Lodhi et al. (1994) proc\ue9dure d\u2019extraction d\u2019acide nucl\ue9ique, en omettant certaines des \ue9tapes sp\ue9cifiques du laboratoire et en faisant la variation de temps d\u2019incubation dans l\u2019azote liquide, au lieu du cong\ue9lateur. L\u2019incubation dans l\u2019azote liquide pendant seulement 1,5 heure a donn\ue9 un ARN de haute qualit\ue9 comme le protocole d\u2019origine, lorsque l\u2019incubation a \ue9t\ue9 effectu\ue9e \ue0 4 \ub0 C pendant une nuit dans un cong\ue9lateur. La transcriptase inverse (RT) a \ue9t\ue9 faite en utilisant un thermocycleur mini PCR portable et l\u2019ADNc, et r\ue9sultant a \ue9t\ue9 amplifi\ue9 en utilisant cette machine mini PCR au lieu d\u2019utiliser un thermocycleur PCR conventionnel stationn\ue9 en laboratoire. L\u2019ADNc a \ue9t\ue9 efficacement amplifi\ue9 et les amplicons \ue9taient similaires \ue0 ceux obtenus avec le protocole d\u2019extraction original et l\u2019amplification ult\ue9rieure par la RT-PCR conventionnelle. Notre protocole a r\ue9duit le temps d\u2019extraction d\u2019environ 16 heures pour le protocole d\u2019origine, \ue0 environ 2 heures et 45 minutes. Si cet outil est utilis\ue9 par le d\ue9partement de la protection des cultures, nous pensons qu\u2019il contribuera grandement \ue0 la production durable de patate douce en faisant des recommandations en temps opportun

EFFECTIVE ISOLATION DISTANCE FOR PREVENTION OF CASSAVA VIRUS INFECTIONS IN UGANDA

Cassava brown streak disease (CBSD) and cassava mosaic disease (CMD) are the major viral diseases of cassava in Uganda. Although isolation distance of \u201c50 m\u201d has been recommended by MAAIF in Uganda for prevention of virus infections in crops, the minimum isolation distance has not been verified for effectiveness in cassava. This study assessed the effective isolation distance for management of viral diseases in cassava. Virus-clean cassava cultivars (NASE 03, NASE 14 and NAROCASS 1) from farmers\u2019 fields were used as field sourced (FS) planting materials. Tissue culture (TC) material of the same cultivars were sourced from the National Crops Resources Research Institute and Makerere University Agricultural Research Institute tissue culture laboratories. Both FS and TC materials were tested at isolation distances of 50, 100, 150 and 250 m for virus prevention. The experiment was laid out in a randomised complete block design and was run for 12 months after planting (MAP). Mean CBSD/CMD prevalence significantly varied (P<0.05) among isolation distances in both FS and TC plants, and the 250 m isolation distance was the most effective in reducing disease prevalence. Across cultivars and planting material category at 12 MAP, the 50 m isolation distance had the highest foliar incidence for CBSD (29.2%) and CMD (16.1%); while severity for CBSD was 1.4 and 1.2 for CMD. At 250 m, all FS and TC plants had CBSD/CMD severity of 1.0 and 0% incidence. These results show that 250 m isolation distance can provide an option to disseminate popular, but CBSD/CMD susceptible cassava cultivars thereby manage CBSD/CMD.La maladie des stries brunes du manioc (CBSD) et la maladie de la mosa\uefque du manioc (CMD) sont les principales maladies virales du manioc en Ouganda. Bien qu\u2019une distance d\u2019isolement de \uab50 m\ubb ait \ue9t\ue9 recommand\ue9e par le MAAIF en Ouganda pour la pr\ue9vention des infections virales dans les cultures, l\u2019efficacit\ue9 minimale de la distance d\u2019isolement n\u2019a pas \ue9t\ue9 v\ue9rifi\ue9e dans le manioc. Cette \ue9tude a \ue9valu\ue9 la distance d\u2019isolement efficace pour la gestion des maladies virales dans le manioc. Des cultivars de manioc sans virus (NASE 03, NASE 14 et NAROCASS 1) provenant des champs des agriculteurs ont \ue9t\ue9 utilis\ue9s comme le source de mat\ue9riel de plantation (FS). Le mat\ue9riel de culture tissulaire (TC) des m\ueames cultivars provenait des laboratoires National Crops Resources Research Institute and Makerere University Agricultural Research Institute tissue culture. Les mat\ue9riaux FS et TC ont \ue9t\ue9 test\ue9s \ue0 des distances d\u2019isolement de 50, 100, 150 et 250 m pour la pr\ue9vention des virus. L\u2019exp\ue9rience a \ue9t\ue9 pr\ue9sent\ue9e dans un bloc complet randomis\ue9 et a \ue9t\ue9 r\ue9alis\ue9e pendant 12 mois apr\ue8s la plantation (MAP). La pr\ue9valence moyenne de CBSD / CMD variait significativement (P <0,05) entre les distances d\u2019isolement dans les plantes FS et TC, et la distance d\u2019isolement de 250 m \ue9tait la plus efficace pour r\ue9duire la pr\ue9valence de la maladie. \uc0 travers les cultivars et la cat\ue9gorie de mat\ue9riel de plantation \ue0 12 MAP, la distance d\u2019isolement de 50 m avait l\u2019incidence foliaire la plus \ue9lev\ue9e pour le CBSD (29,2%) et le CMD (16,1%); tandis que la gravit\ue9 pour CBSD \ue9tait de 1,4 et 1,2 pour CMD. \uc0 250 m, toutes les plantes FS et TC avaient une gravit\ue9 CBSD / CMD de 1,0 et une incidence de 0%. Ces r\ue9sultats montrent qu\u2019une distance d\u2019isolement de 250 m peut fournir une option pour diss\ue9miner des cultivars de manioc sensibles au CBSD / CMD, ce qui permet de g\ue9rer le CBSD / CMD

Pharmacokinetics and pharmacodynamics of azithromycin in severe malaria bacterial co-infection in African children (TABS-PKPD): a protocol for a Phase II randomised controlled trial [version 1; peer review: awaiting peer review]

Background: African children with severe malaria are susceptible to Gram-negative bacterial co-infection, largely non-typhoidal Salmonellae, leading to a substantially higher rates of in-hospital and post-discharge mortality than those without bacteraemia. Current evidence for treating co-infection is lacking, and there is no consensus on the dosage or length of treatment required. We therefore aimed to establish the appropriate dose of oral dispersible azithromycin as an antimicrobial treatment for children with severe malaria and to investigate whether antibiotics can be targeted to those at greatest risk of bacterial co-infection using clinical criteria alone or in combination with rapid diagnostic biomarker tests. Methods: A Phase I/II open-label trial comparing three doses of azithromycin: 10, 15 and 20 mg/kg spanning the lowest to highest mg/kg doses previously demonstrated to be equally effective as parenteral treatment for other salmonellae infection. Children with the highest risk of bacterial infection will receive five days of azithromycin and followed for 90 days. We will generate relevant pharmacokinetic data by sparse sampling during dosing intervals. We will use population pharmacokinetic modelling to determine the optimal azithromycin dose in severe malaria and investigate azithromycin exposure to change in C-reactive protein, a putative marker of sepsis at 72 hours, and microbiological cure (seven-day), alone and as a composite with seven-day survival. We will also evaluate whether a combination of clinical, point-of-care diagnostic tests, and/or biomarkers can accurately identify the sub-group of severe malaria with culture-proven bacteraemia by comparison with a control cohort of children hospitalized with severe malaria at low risk of bacterial co-infection. Discussion: We plan to study azithromycin because of its favourable microbiological spectrum, its inherent antimalarial and immunomodulatory properties and dosing and safety profile. This study will generate new data to inform the design and sample size for definitive Phase III trial evaluation

Pharmacokinetics and pharmacodynamics of azithromycin in severe malaria bacterial co-infection in African children (TABS-PKPD): a protocol for a Phase II randomised controlled trial [version 2; peer review: 1 approved]

Background: African children with severe malaria are susceptible to Gram-negative bacterial co-infection, largely non-typhoidal Salmonellae, leading to a substantially higher rates of in-hospital and post-discharge mortality than those without bacteraemia. Current evidence for treating co-infection is lacking, and there is no consensus on the dosage or length of treatment required. We therefore aimed to establish the appropriate dose of oral dispersible azithromycin as an antimicrobial treatment for children with severe malaria and to investigate whether antibiotics can be targeted to those at greatest risk of bacterial co-infection using clinical criteria alone or in combination with rapid diagnostic biomarker tests. Methods: A Phase I/II open-label trial comparing three doses of azithromycin: 10, 15 and 20 mg/kg spanning the lowest to highest mg/kg doses previously demonstrated to be equally effective as parenteral treatment for other salmonellae infection. Children with the highest risk of bacterial infection will receive five days of azithromycin and followed for 90 days. We will generate relevant pharmacokinetic data by sparse sampling during dosing intervals. We will use population pharmacokinetic modelling to determine the optimal azithromycin dose in severe malaria and investigate azithromycin exposure to change in C-reactive protein, a putative marker of sepsis at 72 hours, and microbiological cure (seven-day), alone and as a composite with seven-day survival. We will also evaluate whether a combination of clinical, point-of-care diagnostic tests, and/or biomarkers can accurately identify the sub-group of severe malaria with culture-proven bacteraemia by comparison with a control cohort of children hospitalized with severe malaria at low risk of bacterial co-infection. Discussion: We plan to study azithromycin because of its favourable microbiological spectrum, its inherent antimalarial and immunomodulatory properties and dosing and safety profile. This study will generate new data to inform the design and sample size for definitive Phase III trial evaluation. Registration: ISRCTN49726849 (27th October 2017)

Characterising demographics, knowledge, practices and clinical care among patients attending sickle cell disease clinics in Eastern Uganda [version 1; peer review: 1 approved, 1 approved with reservations]

Background: In Uganda to date, there are neither established registries nor descriptions of facility-based sickle cell disease (SCD) patient characteristics beyond the central region. Here, we summarize data on the baseline clinical characteristics and routine care available to patients at four clinics in Eastern Uganda as a prelude to a clinical trial. Methods: Between February and August 2018, we conducted a cross-sectional survey of patients attending four SCD clinics in Mbale, Soroti, Atutur and Ngora, all in Eastern Uganda, the planned sites for an upcoming clinical trial (H-PRIME: ISRCTN15724013). Data on socio-demographic characteristics, diagnostic methods, clinic schedules, the use of prophylactic and therapeutic drugs, clinical complications and patient understanding of SCD were collected using a structured questionnaire. Results: Data were collected on 1829 patients. Their ages ranged from 0 to 64 years with a median (IQR) of 6 (3-11) years. 50.9% of participants were male. The majority (1151; 62.9%) reported a positive family history for SCD. Approximately half knew that SCD is inherited from both parents but a substantial proportion did not know how SCD is transmitted and small numbers believed that it is acquired by either transfusion or from other people. Only 118/1819 (6.5%) participants had heard about or were using hydroxyurea while 356/1794 (19.8%) reported stigmatization. Participants reported a median of three (IQR 1-4) hospital admissions during the preceding 12 months; 80.8% had been admitted at least once, while 14.2% had been admitted more than five times. Pain was the most common symptom, while 83.9% of those admitted had received at least one blood transfusion. Conclusion: The majority of patients attending SCD clinics in Eastern Uganda are children and few are currently being treated with hydroxyurea. The data collected through this facility-based survey will provide background data that will be useful in planning for the H-PRIME trial

A predictive algorithm for identifying children with sickle cell anemia among children admitted to hospital with severe anemia in Africa

Sickle cell anemia (SCA) is common in sub-Saharan Africa where approximately 1% of births are affected. Severe anemia is a common cause for hospital admission within the region yet few studies have investigated the contribution made by SCA. The Transfusion and Treatment of severe anemia in African Children Trial (ISRCTN84086586) investigated various treatment strategies in 3983 children admitted with severe anemia (hemoglobin < 6.0 g/dl) based on two severity strata to four hospitals in Africa (three Uganda and one Malawi). Children with known-SCA were excluded from the uncomplicated stratum and capped at 25% in the complicated stratum. All participants were genotyped for SCA at trial completion. SCA was rare in Malawi (six patients overall), so here we focus on the participants recruited in Uganda. We present baseline characteristics by SCA status and propose an algorithm for identifying children with unknown-SCA. Overall, 430 (12%) and 608 (17%) of the 3483 Ugandan participants had known- or unknown-SCA, respectively. Children with SCA were less likely to be malaria-positive and more likely to have an affected sibling, have gross splenomegaly, or to have received a previous blood transfusion. Most outcomes, including mortality and readmission, were better in children with either known or unknown-SCA than non-SCA children. A simple algorithm based on seven admission criteria detected 73% of all children with unknown-SCA with a number needed to test to identify one new SCA case of only two. Our proposed algorithm offers an efficient and cost-effective approach to identifying children with unknown-SCA among all children admitted with severe anemia to African hospitals where screening is not widely available

Randomised controlled trial of oxygen therapy and high-flow nasal therapy in African children with pneumonia.

PURPOSE: The life-saving role of oxygen therapy in African children with severe pneumonia is not yet established. METHODS: The open-label fractional-factorial COAST trial randomised eligible Ugandan and Kenyan children aged > 28 days with severe pneumonia and severe hypoxaemia stratum (SpO2  3 h receipt of oxygen were excluded. The primary endpoint was 48 h mortality; secondary endpoints included mortality or neurocognitive sequelae at 28 days. RESULTS: The trial was stopped early after enrolling 1852/4200 children, including 388 in the severe hypoxaemia stratum (median 7 months; median SpO2 75%) randomised to HFNT (n = 194) or LFO (n = 194) and 1454 in the hypoxaemia stratum (median 9 months; median SpO2 88%) randomised to HFNT (n = 363) vs LFO (n = 364) vs permissive hypoxaemia (n = 727). Per-protocol 15% of patients in the permissive hypoxaemia group received oxygen (when SpO2 < 80%). In the severe hypoxaemia stratum, 48-h mortality was 9.3% for HFNT vs. 13.4% for LFO groups. In the hypoxaemia stratum, 48-h mortality was 1.1% for HFNT vs. 2.5% LFO and 1.4% for permissive hypoxaemia. In the hypoxaemia stratum, adjusted odds ratio for 48-h mortality in liberal vs permissive comparison was 1.16 (0.49-2.74; p = 0.73); HFNT vs LFO comparison was 0.60 (0.33-1.06; p = 0.08). Strata-specific 28 day mortality rates were, respectively: 18.6, 23.4 and 3.3, 4.1, 3.9%. Neurocognitive sequelae were rare. CONCLUSIONS: Respiratory support with HFNT showing potential benefit should prompt further trials

What explains gender inequalities in HIV/AIDS prevalence in sub-Saharan Africa? Evidence from the demographic and health surveys

Abstract Background Women are disproportionally affected by human immunodeficiency virus (HIV)/acquired immunodeficiency syndrome (AIDS) in sub-Saharan Africa (SSA). The determinants of gender inequality in HIV/AIDS may vary across countries and require country-specific interventions to address them. This study aimed to identify the socio-demographic and behavioral characteristics underlying gender inequalities in HIV/AIDS in 21 SSA countries. Methods We applied an extension of the Blinder-Oaxaca decomposition approach to data from Demographic and Health Surveys and AIDS Indicator Surveys to quantify the differences in HIV/AIDS prevalence between women and men attributable to socio-demographic factors, sexual behaviours, and awareness of HIV/AIDS. We decomposed gender inequalities into two components: the percentage attributable to different levels of the risk factors between women and men (the “composition effect”) and the percentage attributable to risk factors having differential effects on HIV/AIDS prevalence in women and men (the “response effect”). Results Descriptive analyses showed that the difference between women and men in HIV/AIDS prevalence varied from a low of 0.68 % (P = 0.008) in Liberia to a high of 11.5 % (P < 0.001) in Swaziland. The decomposition analysis showed that 84 % (P < 0.001) and 92 % (P < 0.001) of the higher prevalence of HIV/AIDS among women in Uganda and Ghana, respectively, was explained by the different distributions of HIV/AIDS risk factors, particularly age at first sex between women and men. In the majority of countries, however, observed gender inequalities in HIV/AIDS were chiefly explained by differences in the responses to risk factors; the differential effects of age, marital status and occupation on prevalence of HIV/AIDS for women and men were among the significant contributors to this component. In Cameroon, Guinea, Malawi and Swaziland, a combination of the composition and response effects explained gender inequalities in HIV/AIDS prevalence. Conclusions The factors that explain gender inequality in HIV/AIDS in SSA vary by country, suggesting that country-specific interventions are needed. Unmeasured factors also contributed substantially to the difference in HIV/AIDS prevalence between women and men, highlighting the need for further study

Elective Cancer Surgery in COVID-19-Free Surgical Pathways During the SARS-CoV-2 Pandemic: An International, Multicenter, Comparative Cohort Study.

PURPOSE: As cancer surgery restarts after the first COVID-19 wave, health care providers urgently require data to determine where elective surgery is best performed. This study aimed to determine whether COVID-19-free surgical pathways were associated with lower postoperative pulmonary complication rates compared with hospitals with no defined pathway. PATIENTS AND METHODS: This international, multicenter cohort study included patients who underwent elective surgery for 10 solid cancer types without preoperative suspicion of SARS-CoV-2. Participating hospitals included patients from local emergence of SARS-CoV-2 until April 19, 2020. At the time of surgery, hospitals were defined as having a COVID-19-free surgical pathway (complete segregation of the operating theater, critical care, and inpatient ward areas) or no defined pathway (incomplete or no segregation, areas shared with patients with COVID-19). The primary outcome was 30-day postoperative pulmonary complications (pneumonia, acute respiratory distress syndrome, unexpected ventilation). RESULTS: Of 9,171 patients from 447 hospitals in 55 countries, 2,481 were operated on in COVID-19-free surgical pathways. Patients who underwent surgery within COVID-19-free surgical pathways were younger with fewer comorbidities than those in hospitals with no defined pathway but with similar proportions of major surgery. After adjustment, pulmonary complication rates were lower with COVID-19-free surgical pathways (2.2% v 4.9%; adjusted odds ratio [aOR], 0.62; 95% CI, 0.44 to 0.86). This was consistent in sensitivity analyses for low-risk patients (American Society of Anesthesiologists grade 1/2), propensity score-matched models, and patients with negative SARS-CoV-2 preoperative tests. The postoperative SARS-CoV-2 infection rate was also lower in COVID-19-free surgical pathways (2.1% v 3.6%; aOR, 0.53; 95% CI, 0.36 to 0.76). CONCLUSION: Within available resources, dedicated COVID-19-free surgical pathways should be established to provide safe elective cancer surgery during current and before future SARS-CoV-2 outbreaks