DNA extraction from silica gel-preserved common bean (Phaseolus vulgaris L.) leaves

Extraction of non-degraded and contaminant-free DNA from field specimen requires collection under liquid nitrogen which is not readily available in resource constrained laboratories in low and middle income countries (LMICs). A method of extracting DNA from silica gel-preserved common bean (Proteus vulgaris L.) leaves is presented. The method, which does not involve the use of phenol, chloroform or isoamyl alcohol also obviates the need for low temperature incubation during the DNA extraction steps and the grinding of desiccated leaf tissue in liquid nitrogen. It relies on inactivating proteins using SDS and proteinase K along with precipitation of polysaccharides using a high salt solution (0.8 M NaCl). DNA is further purified by exploiting its insolubility in aqueous media. High quality pure DNA (mean concentration 2.84 ± 0.013 µg/g of dry leaf tissue) with mean DNA purity values of 2.1 ± 0.1 was extracted. The DNA was also found to be free of protein and polysaccharide contamination. This method enables DNA amplification using molecular markers routinely used in molecular biology laboratories like random amplified polymorphic (RAPD) markers, inter simple sequence repeat (ISSR) markers, sequence-characterized amplified region (SCAR) markers and simple sequence repeat (SSR) markers. The findings of this study show that it is possible to obtain high quality DNA from leaf tissue preserved in silica gel. The method used in this research will be invaluable to resource constrained laboratories in low and middle income countries (LMICs) that cannot afford to buy or access liquid nitrogen in order to extract high quality DNA and for research groups undertaking field surveys that require several days or weeks off station without laboratory freezers to maintain the integrity of the tissues which is crucial for obtaining high quality DNA

A simple and efficient method for extracting S. Rolfsii DNA for PCR based diversity studies

Present methods of extracting DNA from Sclerotium rolfsii use a lot of hazardous organic chemicals to extract high quality DNA. Extraction of the DNA is further complicated by exopolysaccharides that bind to the DNA making it mucilaginous. We developed a simple and efficient protocol for extracting DNA high quality from S. rolfsii. Our method uses a DNA extraction buffer that contains sodium dodecyl sulphate and proteinase K to inactivate proteins and high salt concentration to precipitate the exopolysaccharides. It uses neither phenol, chloroform nor isoamyl alcohol during the DNA extraction process. It also does not require freeze drying of the mycelia and grinding in liquid nitrogen. Using our method, a sufficient amount of pure (mean A260: A280=1.91 ± 0.001) DNA (mean = 55.57 ± 0.002 ng/µl) was obtained from 100 mg of mycelia. The DNA was amenable to PCR amplification using inter-simple sequence repeat primers and primers targeting the internal transcribed spacer region of S. rolfsii. Our method will be very useful in laboratories that don’t have access to liquid nitrogen and freeze-drying facilities and will be a catalyst for PCR-based phylogenetic studies of this important pathogen of common bean

Effectiveness of pyramided genes in conferring resistance to anthracnose disease in common bean populations

Anthracnose disease (Colletotrichum lindemuthianum (Sacc. et. Magn) Lams. Scrib.) is one of the most devastating diseases that constrain common bean production in Uganda. A cascading pedigree pyramiding scheme was used to develop common bean populations to evaluate the effectiveness of pyramided and single resistance genes (Co-42, Co-43, Co-5, and Co-9) on disease development. Detached leaf trifoliates of F4:6 genotypes were screened with four C. lindemuthianum races (352, 713,767 and 2047). Disease severity data were subjected to ANOVA. Races, genotypes and Race x Genotype interaction were significant. Genes Co-42 and Co-5 conferred resistance to the four races and the gene pyramids Co-42+Co-5+Co-9 and Co-42+Co-5 had the lowest severity scores. Gene Co-43 conferred resistance to race 352 and weak resistance to race 713; whereas gene Co-9 conferred resistance to race 352. Co-43+Co-9 gene pyramid showed resistance only to race 352. The Co-42 and Co-5 genes conferred resistance to all the four races 352, 713, 767 and 2047. The single gene Co-42 was not significantly different from the pyramids Co-42+Co-5+Co-9 and Co-42+Co-5 (P<0.01). Similarly, the Co-5gene was not significantly different from Co-42+Co-5, Co-42+Co-9 and Co-5+Co-9 pyramids. The Co-9gene showed antagonism in all pyramids. These results indicate that pyramiding of resistance genes would be effective for disease management in Uganda, but pyramids with Co-9 gene would be less effective

Development of common bean genotypes with high iron and zinc seed concentrations and superior canning and agronomic quality traits

Iron deficiency anemia is prevalent worldwide but mainly affects children under five years of age and women of reproductive age. One of the main causes of anemia in these groups is diet incapable of meeting daily iron requirements. Biofortification of staple foods is an approach aimed at contributing to reduction of anemia in Africa, and common bean (Phaseolus vulgaris L.), one of the leading staple foods in East and Central Africa, has gained attention as a valuable source of iron (Fe) and zinc (Zn). Its usefulness in improving iron status of iron anemic women and children is documented. Natural variation in iron and associated micronutrients like zinc exists in beans but their concentrations are below the target levels to meet daily requirements. This study aimed to develop and identify potential bean genotypes that surpass the HarvestPlus threshold of 90 ppm seed iron for possible promotion as high iron and zinc beans, and utilization in hybridization programs targeting these minerals, productivity and market traits. Advanced 578 genotypes were evaluated in five genotype groups planted in three locations from 2016 to 2018. Genotypes significantly differed (P≤0.05) in Fe, Zn, cooking time, canning quality and yield. Iron and Zn varied highly, ranging between 44-118 and 25-50 ppm, respectively, across the five-genotype groups. Cooking time ranged from 29-118 minutes and majority of the genotypes expressed good to excellent canning quality based on visual assessment (4-5) and hydration coefficients (2.1-2.2). Mean yields for bush beans and climbers were 1674-1977 kg ha-1 and 2204-3160 kg ha-1, respectively. The most outstanding genotypes that combined above average yield with high Fe/Zn were CMKN1109 (96/ 43 ppm), SMR103 (92/ 43 ppm), SMC12 (90/43 ppm), and NUS16 (91/ 48 ppm). In addition, NUA127 (84/ 42 ppm), SMR53 (84/42 ppm), SMC160 (84/ 43 ppm) and NUA595 (83/ 42 ppm) yielded above average and expressed high canning quality. The genotypes that combined high Fe/ Zn, canning quality, and yield are potential genotypes for further improvement or evaluation for possible release

Analysis of genetic diversity of banana weevils (Cosmopolites sordidus) (Coleoptera: Curculionidae) using tanscriptome-derived simple sequence repeat markers

The banana weevil, Cosmopolites sordidus (Germar) (Coleoptera: Curculionidae) is an economically important insect pest of bananas. It causes up to 100% yield losses and substantial lifespan reduction in bananas. Advances in genomics, proteomics, and sequencing technologies have provided powerful pathways to genotyping disastrous pests such as C. sordidus. However, such technologies are often not available to the majority of rural subtropical African banana growers and pest control managers. This study was therefore motivated by the need to create cheap and easily accessible C. sordidus genotyping methods that could be deployed by banana pest control managers to the benefit of C. sordidus control programs in the tropics where such advanced technologies are not readily accessible. We used an in-house C. sordidus transcriptome from the an-ongoing study from which we mined an array of simple sequence repeat (SSR) markers. Of these, six highly polymorphic transcriptome-derived SSR markers were used to successfully genotype within and among banana weevil population genetic diversity of 12 C. sordidus populations collected from four banana-growing agro-ecological zones (AEZs) in Uganda. The developed transcriptome-derived SSR markers can be used by researchers in population genetics for characterization of the C. sordidus and identification of new genes that are linked to traits of particular interest. The significant genetic diversity revealed in C. sordidus provides pertinent information for integrated pest management strategies

Response of common bean genotypes to prevalent Pseudocercospora griseola races causing angular leaf spot in Uganda

Angular Leaf Spot (ALS) caused by Pseudocercospora griseola is responsible for 54% yield loss in Uganda’s common beans. Host plant resistance is a safe and cost-effective management strategy for this disease. Identification of resistant common bean genotypes to prevailing races is vital to utilize the crop. Therefore, the objective of this study was to identify genotypes that are resistant to the commonly occurring and virulent P. griseola races in Uganda for breeding purposes. Twenty-seven common bean genotypes and three control varieties (Mexico54, MCM5001, and CAL96) were screened at field conditions for ALS resistance at testing site (National Agricultural Research Laboratories - Kawanda) under natural disease infection. The genotypes were also evaluated in the screen house using frequently occurring P. griseola races: 61:63, 1:6 and 21:39. Variability in the severity of ALS on both leaves and pods was significant whereas the difference between seasons and the interaction between the seasons and genotypes was only significant for yield. The disease severity scores were higher (mean of 3.2) on leaves than on pods (mean of 2.9). Ninety-three percent, 33.3% and 15% of the genotypes were resistant to P. griseola races 21:39, 1:6 and 61:63, respectively. A large-seeded genotype AFR703 was resistant to all the three P. griseola races. A medium seed size genotype AFR702 and three small seed genotypes (G148, G18842 and G6727) were resistant to both 21:39 and 1:6 but moderate resistance to 61:63 whereas a large-seeded genotype AND279 was resistant to both 61:63 and 21:39 but moderate to 1:6. All of these six genotypes (AFR703, AFR702, G148, G18842, G6727 and AND279) expressed moderate resistance to P. griseola races on leaves under field conditions. Thus, these common bean genotypes could be used as sources of ALS resistance for breeding programs to address the ALS constraint; and genes responsible for resistance have to be characterized. Key words: Pathology, Disease, Resistance, Infection, BALSIT, Pathotypes, Prevalent, Race

Gain and performance in yield and micronutrient concentration in common bean improvement

Common bean (Phaseolus vulgaris L.) is a staple and nutritious leguminous food crop for all income categories in Africa. Efforts to improve its yield performance and nutritional components, especially Iron and zinc have resulted in the release of several varieties in the sub-Saharan African region. The objective of this study was to assess genetic progress in varieties released in 12 African countries through the Pan African Bean Research Alliance (PABRA) from 1973 to 2017, to inform current breeding decisions. A total of 214 released varieties, land races and breeding lines, of bush and climbing beans were evaluated for yield, micronutrient (Fe and Zn) concentration, and phenology in three locations (Kawanda and Kachwekano in Uganda, and Kitengule in Tanzania) in 2017–2018. There were significant differences (P 0.01) among genotypes for all traits except days to maturity (DPM). Genotype x environment interaction was also significant (P<0.05) for all assessed traits, except for iron in climbing beans. Across environments, repeatability (H2) was greater than 0.50 for all traits, except for DPM in climbing beans. Annual rates of genetic yield gains were 4.41 and 4.57 kg ha-1 for large and small seed bush beans, and -2.74 and 21.6 kg ha-1 for large and small seeded climbers. Similarly, gains in seed iron (FESEED) were 0.40 and 0.17 ppm for bush and climbing beans, respectively. These represented an annual relative gain over the oldest varieties of 0.6 and 0.7% kg ha-1 for yield of large and small seeded bush beans, -0.3 and 1.6% kg ha-1 for yield of large and small seeded climbers, 0.6 and 0.2% ppm for FESEED of bush and climbers. Overall, genetic progress was slow for both yield and FESEED

Co-evolution of NK receptors and HLA ligands in humans is driven by reproduction.

Allogeneic individuals co-exist during pregnancy in eutherian mammals. Maternal and fetal cells intermingle at the site of placental attachment in the uterus, where the arteries are remodeled to supply the fetus with oxygen and nutrients. This access by placental cells to the maternal supply line determines the growth and birth weight of the baby and is subject to stabilizing selection. Invading placental trophoblast cells express human leukocyte antigen class I ligands (HLA-E, HLA-G, and HLA-C) for receptors on maternal uterine natural killer (NK) and myelomonocytic cells, CD94/NKG2, leukocyte immunoglobulin-like receptor (LILR), and killer immunoglobulin receptor (KIR). Of these, only the KIR/HLA-C system is highly polymorphic. Different combinations of maternal KIR and fetal HLA-C variants are correlated with low birth weight and pre-eclampsia or high birth weight and obstructed labor, the two extremes of the obstetric dilemma. This situation has arisen because of the evolution of bipedalism and subsequently, in the last million years, larger brains. At this point, the human system began to reach a balance between KIR A and KIR B haplotypes and C1 and C2 epitopes of HLA-C alleles that reflects a functional compromise between the competing demands of immunity and reproduction.We are grateful for financial support from Centre for Trophoblast Research, University of Cambridge, King's College, Cambridge, and the Wellcome Trust.This is the author accepted manuscript. The final version is available from Wiley via http://dx.doi.org/10.1111/imr.1232

Increased Avian Diversity Is Associated with Lower Incidence of Human West Nile Infection: Observation of the Dilution Effect

Recent infectious disease models illustrate a suite of mechanisms that can result in lower incidence of disease in areas of higher disease host diversity–the ‘dilution effect’. These models are particularly applicable to human zoonoses, which are infectious diseases of wildlife that spill over into human populations. As many recent emerging infectious diseases are zoonoses, the mechanisms that underlie the ‘dilution effect’ are potentially widely applicable and could contribute greatly to our understanding of a suite of diseases. The dilution effect has largely been observed in the context of Lyme disease and the predictions of the underlying models have rarely been examined for other infectious diseases on a broad geographic scale. Here, we explored whether the dilution effect can be observed in the relationship between the incidence of human West Nile virus (WNV) infection and bird (host) diversity in the eastern US. We constructed a novel geospatial contrasts analysis that compares the small differences in avian diversity of neighboring US counties (where one county reported human cases of WNV and the other reported no cases) with associated between-county differences in human disease. We also controlled for confounding factors of climate, regional variation in mosquito vector type, urbanization, and human socioeconomic factors that are all likely to affect human disease incidence. We found there is lower incidence of human WNV in eastern US counties that have greater avian (viral host) diversity. This pattern exists when examining diversity-disease relationships both before WNV reached the US (in 1998) and once the epidemic was underway (in 2002). The robust disease-diversity relationships confirm that the dilution effect can be observed in another emerging infectious disease and illustrate an important ecosystem service provided by biodiversity, further supporting the growing view that protecting biodiversity should be considered in public health and safety plans

Psychosocial impact of undergoing prostate cancer screening for men with BRCA1 or BRCA2 mutations.

OBJECTIVES: To report the baseline results of a longitudinal psychosocial study that forms part of the IMPACT study, a multi-national investigation of targeted prostate cancer (PCa) screening among men with a known pathogenic germline mutation in the BRCA1 or BRCA2 genes. PARTICPANTS AND METHODS: Men enrolled in the IMPACT study were invited to complete a questionnaire at collaborating sites prior to each annual screening visit. The questionnaire included sociodemographic characteristics and the following measures: the Hospital Anxiety and Depression Scale (HADS), Impact of Event Scale (IES), 36-item short-form health survey (SF-36), Memorial Anxiety Scale for Prostate Cancer, Cancer Worry Scale-Revised, risk perception and knowledge. The results of the baseline questionnaire are presented. RESULTS: A total of 432 men completed questionnaires: 98 and 160 had mutations in BRCA1 and BRCA2 genes, respectively, and 174 were controls (familial mutation negative). Participants' perception of PCa risk was influenced by genetic status. Knowledge levels were high and unrelated to genetic status. Mean scores for the HADS and SF-36 were within reported general population norms and mean IES scores were within normal range. IES mean intrusion and avoidance scores were significantly higher in BRCA1/BRCA2 carriers than in controls and were higher in men with increased PCa risk perception. At the multivariate level, risk perception contributed more significantly to variance in IES scores than genetic status. CONCLUSION: This is the first study to report the psychosocial profile of men with BRCA1/BRCA2 mutations undergoing PCa screening. No clinically concerning levels of general or cancer-specific distress or poor quality of life were detected in the cohort as a whole. A small subset of participants reported higher levels of distress, suggesting the need for healthcare professionals offering PCa screening to identify these risk factors and offer additional information and support to men seeking PCa screening