Eco-Geographical and Botanical Patterns of Resistance to Lepidoptera Insects in Brassica rapa L.

In the context of the widespread expansion of damage by herbivorous pests of Brassica crops, taking into account the requirements for minimizing pesticide pollution of the environment, it is important to have fundamental knowledge of the geographical features of the distribution of pests and about the botanical confinement of plant resistance in order to develop a strategy for creating new Brassica cultivars with complex resistance to insects. The relevance of our work is related to the study of the variability in the degree of resistance of the extensive genetic diversity of Brassica rapa accessions to the main herbivorous pests of Brassica crops in contrasting ecological and geographical zones of the Russian Federation (Arctic, northwestern, and southern zones). We have studied the distribution and food preferences of Lepidoptera insects (diamondback moth Plutella xylostella and cabbage moth Mamestra brassicae) on a set of 100 accessions from the VIR B. rapa collection (Chinese cabbage, pakchoi, wutacai, zicaitai, mizuna, and leaf and root turnips) in the field in three zones of the Russian Federation. We have found that the diamondback moth and cabbage moth are largely harmful in three zones of the European part of the Russian Federation, although the degree of damage to plants by these insects varies by year of cultivation. On average, for the set studied during the two years of the experiment, the degree of plant damage by both pests in the Arctic zone was low and almost low, and in the northwestern and southern zones, it was medium. It was noted that diamondback moth damage was greater in the northwestern zone in both years and in the southern and Arctic zones in 2021, while in 2022, the degree of cabbage moth damage was slightly higher in the southern and Arctic zones. Under the conditions of field diamondback moth damage, the accessions of Chinese cabbage, wutacai, and mizuna turned out to be the most resistant (the damage score was 1.92-1.99), whereas the accessions of wutacai and pakchoi were the most resistant to the cabbage moth (the damage score was 1.62-1.78). A high variability in the degree of resistance of Brassica crops to Lepidoptera insects from complete resistance to susceptibility was revealed. We have identified sources of resistance to insects, including complex resistance in all study areas, among landraces and some modern cultivars of Chinese cabbage, pakchoi, wutacai, and mizuna from Japan and China, as well as European turnips. The highest susceptibility to pests in the studied set was noted in the accession of root turnip "Hinona" (k-1422, USA) (average damage score of 3.24-3.53 points). We were not able to establish the morphological features of resistant plants or the geographical confinement of the origin of resistance of B. rapa crop accessions.

Resistance of breast cancer cells to paclitaxel is associated with low expressions of miRNA-186 and miRNA-7.

Aim: Neo-adjuvant chemotherapy is a common approach for the complex treatment of breast cancer (BC) and paclitaxel (PTX) is frequently included in the therapeutic regimen. However, the effect of PTX-based treatment is hard to predict precisely based on routinely used markers. As microRNAs are considered a new promising class of biomarkers, the link between miRNA expression and PTX resistance of BC cells needs to be well investigated. This study aimed at the identification of miRNAs associated with responses of BC cells to PTX. Methods: Intrinsic PTX sensitivity and miRNA profiling were assayed in five BC cell lines to identify candidate miRNAs. Selected miRNA (n. 15) expressions were analyzed by real-time-quantitative polymerase chain reaction (RT-qPCR) in BC tissue samples (n. 31) obtained from a diagnostic biopsy. Results were analyzed in the context of the effect of two cycles of PTX and the effect of the completed scheme of neoadjuvant therapy. The study's design facilitated the evaluation of the effect of PTX on cells and the identification of features of the microRNA expression profiles associated exclusively with sensitivity to this drug. Results: miR-186 and miR-7 expression in BC tissues was higher in patients with better outcomes of PTX-based neoadjuvant therapy. Conclusion: High expressions of miR-186 and miR-7 are associated with good response to PTX, whereas their low expressions may be associated with resistance to PTX in BC, indicating the possibility of developing innovative test systems for the prediction of the PTX response, which can be used before the start of neo-adjuvant chemotherapy for BC.

Melon diversity on the Silk Road by molecular phylogenetic analysis in Kazakhstan melons.

To uncover population structure, phylogenetic relationship, and diversity in melons along the famous Silk Road, a seed size measurement and a phylogenetic analysis using five chloroplast genome markers, 17 RAPD markers and 11 SSR markers were conducted for 87 Kazakh melon accessions with reference accessions. Kazakh melon accessions had large seed with exception of two accessions of weedy melon, Group Agrestis, and consisted of three cytoplasm types, of which Ib-1/-2 and Ib-3 were dominant in Kazakhstan and nearby areas such as northwestern China, Central Asia and Russia. Molecular phylogeny showed that two unique genetic groups, STIa-2 with Ib-1/-2 cytoplasm and STIa-1 with Ib-3 cytoplasm, and one admixed group, STIAD combined with STIa and STIb, were prevalent across all Kazakh melon groups. STIAD melons that phylogenetically overlapped with STIa-1 and STIa-2 melons were frequent in the eastern Silk Road region, including Kazakhstan. Evidently, a small population contributed to melon development and variation in the eastern Silk Road. Conscious preservation of fruit traits specific to Kazakh melon groups is thought to play a role in the conservation of Kazakh melon genetic variation during melon production, where hybrid progenies were generated through open pollination.

The significance of upfront autologous stem cell transplantation for high-intermediate/high-risk stage IV diffuse large B-cell lymphoma.

BACKGROUND: Diffuse large B-cell lymphoma (DLBCL) is the most common (30%-35%) type of B-cell lymphoma. Only about 60% of all newly diagnosed advanced-stage DLBCL can be completely treated with x6 R-CHOP. High-dose chemotherapy (HDCT) followed by autologous hematopoietic stem cell transplantation in the first remission (upfront auto-HSCT) can serve as an option to improve a prognosis in these patients. AIMS: This trial aimed to improve prognosis in DLBCL by upfront auto-HSCT. METHODS AND RESULTS: A group of 105 patients: DLBCL NOS, age 18-65, stage IV, IPI ≥2, CR/PR after x6 R-CHOP/DA-EPOCH-R from 2010 to 2019 at NMRC of Oncology named after N.N.Petrov of MoH of Russia was retrospectively analyzed. The HSCT group included patients with upfront HDCT followed by auto-HSCT (n = 35). The control group included patients with non-invasive follow-up after induction (n = 70). Primary endpoint was progression-free survival (PFS). Secondary endpoints were overall survival (OS), response rate and relapse rate. The 3-year OS (p = .013) and 3-year PFS (p = .033) were significantly higher in the HSCT group. The 3-year OS was decreased by the occurrence of relapse (p ≤ .001) and weight loss (B-symptom) (p = .04). DEL was the negative prognostic factor for 3-year PFS in all patients (p = .001) and control group (p = .001). DA-EPOCH-R significantly increased the 3-year PFS (p = .041). CONCLUSION: Upfront HDCT followed by auto-HSCT can increase 3-year OS and PFS and improve prognosis in DLBCL NOS, age 18-65, stage IV, IPI ≥2 patients.

An MSH6 germline pathogenic variant p.Gly162Ter associated with Lynch syndrome.

We identified a three-generation Russian family with Lynch syndrome with a novel germline variant of the MSH6 gene. An 84-year-old female was diagnosed with endometrial adenocarcinoma at the age of 49 years. Her son was diagnosed with colorectal tubular adenoma at the age of 32 years. A germline nonsense variant (c.484 G > T:p.Gly162Ter) in exon 3 of the MSH6 gene was revealed by whole-exome sequencing. Sanger sequencing confirmed the cosegregation of the MSH6 nonsense variant in family members.

Immunology and immunotherapy in breast cancer.

Immuno-oncology is a rapidly developing field in medicine. Drug combination therapies have already been studied in many clinical trials on various tumor types. In recent years, a checkpoint inhibition therapy with monoclonal antibodies targeting PD-1 and its ligand PD-L1 has been developed. Breast cancer had been examined in the field of immune-oncology relatively recently. This review focuses on clinical evidence regarding immune checkpoint inhibition for curative treatment of various breast cancer subtypes. In addition, we present the results of studies demonstrating the prognostic and predictive value of levels of tumorinfiltrating lymphocytes (CD4 and CD8), their quantitative ratios, and their correlation with regulatory genes (PD-1, PD-L1, and FOX-P3).

Single photon emission computed tomography-computed tomography visualization of sentinel lymph nodes for lymph flow guided nodal irradiation in oral tongue cancer.

PURPOSE: To evaluate correlation of single photon emission computed tomography-computed tomography (SPECT-CT) data on lymph flow (LF) from oral tongue cancer (OC) and the topography of lymph nodes (LN) metastases; to determine the clinical value of lymph flow guided radiotherapy (LFGRT). MATERIALS AND METHODS: SPECT-CT visualization of LF from the OC lesions was performed after peritumoral injection of 99mTc-phytate in 26 primary patients with clinical stage cT1-2N0M0 disease. We determined the individual drainage (unilateral/bilateral) from the tumor, and localization of sentinel LNs according to the neck levels. Metastases in LNs were verified with histology and a 2-year follow-up. RESULTS: SPECT-CT detected bilateral LF in 10 (38.5%) of 26 patients; in 16 (61.5%) cases the drainage was unilateral. Histology revealed LNs metastases in three cases; regional recurrences were diagnosed in other four patients. In all seven observations metastases were located at the same site and level as the sentinel LNs. In eight (30.8%) of 26 patients sentinel LNs were visualized unilaterally at levels Ib-IIa; in five cases, unilaterally at levels I-IIa-III. In these patients, LFGRT demonstrated 59%-70% reduction of irradiated volume, and 26%-42% and 51%-70% decrease of the mean dose to the spinal cord and the contralateral parotid gland. In patients with a bilateral drainage the reduction of doses absorbed by the spinal cord and contralateral parotid gland was 19% and 6%, respectively. CONCLUSION: Localization of sentinel LNs determined by SPECT-CT corresponds to the localization of metastatic LNs in terms of side and levels.

Genetic Diversity of Phenotypic and Biochemical Traits in VIR Radish (Raphanus sativus L.) Germplasm Collection.

Small radish and radish are economically important root crops that represent an integral part of a healthy human diet. The world collection of Raphanus L. root crops, maintained in the VIR genebank, includes 2810 accessions from 75 countries around the world, of which 2800 (1600 small radish, 1200 radish) belong to R. sativus species, three to R. raphanistrum, three to R. landra, and four to R. caudatus. It is necessary to systematically investigate the historical and modern gene pool of root-bearing plants of R. sativus and provide new material for breeding. The material for our research was a set of small radish and radish accessions of various ecological groups and different geographical origin, fully covering the diversity of the species. The small radish subset included 149 accessions from 37 countries, belonging to 13 types of seven varieties of European and Chinese subspecies. The radish subset included 129 accessions from 21 countries, belonging to 18 types of 11 varieties of European, Chinese, and Japanese subspecies. As a result of the evaluation of R. sativus accessions according to phenological, morphological, and biochemical analyses, a wide variation of these characteristics was revealed, which is due to the large genetic diversity of small radish and radish of various ecological and geographical origins. The investigation of the degree of variation regarding phenotypic and biochemical traits revealed adaptive stable and highly variable characteristics of R. sativus accessions. Such insights are crucial for the establishment and further use of trait collections. Trait collections facilitate germplasm use and contribute significantly to the preservation of genetic diversity of the gene pool.

Opening the Treasure Chest: The Current Status of Research on Brassica oleracea and B. rapa Vegetables From ex situ Germplasm Collections.

Germplasm collections reflect the genetic variability in crops and their wild relatives. Hence, those genetic resources are tremendously valuable for breeders and researchers, especially in light of climatic change and stagnant crop production rates. In order to achieve improvements in crop production and end-use quality, favorable traits and donor alleles present in germplasm collections need to be identified and utilized. This review covers recent reports on the utilization of germplasm material to isolate genotypes of Brassica oleracea and B. rapa vegetables, focusing on high nutrient use efficiency, accumulation of biologically active metabolites, pest resistance, and favorable phenotypic appearance. We discuss the current state of Brassica vegetable collections in genebanks and summarize studies directed to the molecular characterization of those collections.

Myocardial PD-L1 Expression in Patients With Ischemic and Non-ischemic Heart Failure.

Objective: Immune checkpoints inhibitors are promising and wide-spread agents in anti-cancer therapy. However, despite their efficacy, these agents could cause cardiotoxicity, a rare but life-threatening event. In addition, there are still no well-described predictive factors for the development of immune-related adverse events and information on high risk groups. According to known experimental studies we hypothesized that cardiovascular diseases may increase myocardial PD-L1 expression, which could be an extra target for Checkpoint inhibitors and a potential basis for complications development. Methods: We studied patterns of myocardial PD-L1 expression in non-cancer-related cardiovascular diseases, particularly ischemic heart disease (n = 12) and dilated cardiomyopathy (n = 7), compared to patients without known cardiovascular diseases (n = 10) using mouse monoclonal anti-PD-L1 antibody (clone 22C3, 1:50, Dako). Correlation between immunohistochemical data and echocardiographic parameters was assessed. Statistical analyses were performed using R Statistical Software-R studio version 1.3.1093. Results: In the myocardium of cardiac patients, we found membranous, cytoplasmic, and endothelial expression of PD-L1 compared to control group. In samples from patients with a history of myocardial infarction, PD-L1 membrane and endothelial expression was more prominent and frequent, and cytoplasmic and intercalated discs staining was more localized. In contrast, samples from patients with dilated cardiomyopathy displayed very faint endothelial staining, negative membrane staining, and more diffuse PD-L1 expression in the cytoplasm and intercalated discs. In samples from the non-cardiac patients, no convincing PD-L1 expression was observed. Moreover, we discovered a significant negative correlation between PD-L1 expression level and left ventricular ejection fraction and a positive correlation between PD-L1 expression level and left ventricular end-diastolic volume. Conclusions: The present findings lay the groundwork for future experimental and clinical studies of the role of the PD-1/PD-L1 pathway in cardiovascular diseases. Further studies are required to find patients at potentially high risk of cardiovascular adverse events associated with immune checkpoint inhibitors therapy.

Melatonin Administered before or after a Cytotoxic Drug Increases Mammary Cancer Stabilization Rates in HER2/Neu Mice.

INTRODUCTION: The effects of chemotherapy are known to depend on the time of administration. Circadian rhythms are disturbed in tumors and in tumor bearers. Agents involved in controlling the circadian rhythms (chronobiotics) potentially can modify the outcomes of chemotherapeutics administered at different times of the day. Pineal hormone melatonin (MT) is a prototypic chronobiotic. OBJECTIVE: The aim of the study was to investigate if MT can affect efficacy or toxicity of chemotherapy drugs administered at the extreme time points of the working day of hospital personnel. METHODS: Cyclophosphamide, adriamycin, and 5-fluorouracil (CAF) and adriamycin and docetaxel (AT) cytotoxic drug combinations were administered on day 0 at 11:00 a.m. or at 5:00 p.m. (UTC+03:00) to 6-month-old female HER2/neu transgenic FVB/N mice bearing mammary adenocarcinomas. Some mice were additionally provided with MT in drinking water (20 mg/L) at night 1 week before or 3 weeks after treatment or during both periods. Tumor node sizes, body weight, and blood cell counts were determined right before treatment and on days 2, 7, 14, and 21. RESULTS: Significant decrease in the mean tumor node volume was found by days 14 and 21 upon all CAF and AT treatment schedules, except in animals treated with AT at 5:00 p.m. without supplementation with MT. In the latter case, mean tumor node volume on day 21 was the same as in the control. Supplementation of AT administered at 5:00 p.m. with MT improved the tumor response. CAF and AT regimens supplemented with MT also augmented the number of tumor nodes that did not increase by more than 20% by day 21 as compared to CAF or AT alone, respectively. This effect was significant in groups treated with AT at 5:00 p.m. and consistent upon other schedules. On day 7, leukopenia and anemia were registered in groups treated with CAF regimen; however, blood cell counts normalized by day 14. Both CAF and AT were associated with drop in the body weight registered on day 7. Supplementation with MT did not affect changes of the body weight and blood counts. CONCLUSIONS: MT supplementation to cytotoxic drugs can improve antitumor response, especially if it is blunted because of an inappropriate time of administration.

SPECT-CT visualization and biopsy of sentinel lymph nodes in patients with stage IAB-IIA cervical cancer.

OBJECTIVE: To evaluate the diagnostic accuracy of sentinel lymph node (SLN) biopsy and to determine clinical significance of preoperative single-photon emission computed tomography/computed tomography (SPECT-CT). METHODS: We retrospectively evaluate data and images of 55 women with stage IAB-IIA cervical cancer that underwent sentinel lymph node biopsy and subsequent lymph node dissection. 99mTc-phytate (300 MBq) was injected submucosally in 4 points and SPECT-CT started 2 h after the injection. All lymph nodes (LN) that accumulated radiocolloids were regarded as sentinel. Next day after the mapping radical hysterectomy with complete pelvic and parametrial LN dissection and SLN biopsy were performed according to the institute protocol. RESULTS: SPECT-CT identified SLNs in 51 of 55 patients. All SLNs were successfully detected during the surgery. The SLN mapping efficacy was 92.7%. LN metastases were diagnosed in 6/51 (11.8%) cases, including 2 patients with positive SLNs. Sensitivity, negative predictive value (NPV) and false-negative rate for SLN biopsy were as follows: 33.4%, 91.8% and 66.6%. In patients with unilateral SPECT-CT localization of SLNs, LN metastases were identified in 4 (20%) of 20 patients but SLN biopsy was false negative in all cases. In 31 women, SPECT-CT detected SLNs on both sides of the pelvis. In this group, regional LN metastases were diagnosed in 2 patients, and in both cases metastases were detected only in SLNs. CONCLUSIONS: In women with SPECT-CT visualization of SLNs on both sides of the pelvis, SLN biopsy had high sensitivity and NPV. In patients with unilateral SLN localization, SLN biopsy cannot be used for the prediction of regional LN status.

SNP Markers and Evaluation of Duplicate Holdings of Brassica oleracea in Two European Genebanks.

Around the world, there are more than 1500 genebanks storing plant genetic resources to be used in breeding and research. Such resources are essential for future food security, but many genebanks experience backlogs in their conservation work, often combined with limited budgets. Therefore, avoiding duplicate holdings is on the agenda. A process of coordination has started, aiming at sharing the responsibility of maintaining the unique accessions while allowing access according to the international treaty for plant genetic resources. Identifying duplicate holdings based on passport data has been one component of this. In the past, and especially in vegetables, different selections within the same varieties were common and the naming practices of cultivars/selections were flexible. Here, we examined 10 accession pairs/groups of cabbage (Brassica oleracea var. capitata) with similar names maintained in the Russian and Nordic genebanks. The accessions were analyzed for 11 morphological traits and with a SNP (Single Nucleotide Polymorphism) array developed for B. napus. Both proved to be useful tools for understanding the genetic structure among the accessions and for identifying duplicates, and a subset of 500 SNP markers are suggested for future Brassica oleracea genetic characterization. Within five out of 10 pairs/groups, we detected clear genetic differences among the accessions, and three of these were confirmed by significant differences in one or several morphological traits. In one case, a white cabbage and a red cabbage had similar accession names. The study highlights the necessity to be careful when identifying duplicate accessions based solely on the name, especially in older cross-pollinated species such as cabbage.

Effects of Genotype and Culture Conditions on Microspore Embryogenesis and Plant Regeneration in Brassica Rapa ssp. Rapa L.

Turnip is a biennial crop and, consequently, the creation of pure lines for breeding is a time-consuming process. The production of pure turnip lines using doubled haploids produced in isolated microspore culture has not been sufficiently developed. The aim of the present work was to determine some key factors inducing embryogenesis in the isolated microspore culture of turnip, as well as investigating the manners of embryo development. It was shown that the acidity of the medium is an important factor in embryo production; different optimal pH levels ranging from 6.2 to 6.6 corresponded to individual genotypes. Such factors as the cold treatment of buds and the addition of activated charcoal to the nutrient medium increased the responsiveness of all genotypes studied. The turnip variety 'Ronde witte roodkop herfst' demonstrated a genetic disorder in the development of microspores; namely, non-separation of some microspores from tetrads. In the in vitro culture, each of the daughter microspores developed on its own. This indicates the dependence of the possibility of embryogenesis in the turnip microspore culture on the genotype. Results suggest that the initiation of secondary embryogenesis in primary embryos leads to an increase in the proportion of doubled haploid plants.

99mTc-MIBI scintimammography and digital mammography in the diagnosis of multicentric breast cancer.

OBJECTIVE: To compare diagnostic accuracy of scintimammography (SMG) and digital mammography (DS) in the diagnosis of multicentric breast cancer. SUBJECTS AND METHODS: Three hundred and sixty seven women with histologically confirmed breast cancer were included in this analysis. All patients were candidates for conservative breast surgery and had cT1-3N0-1 stage of disease. Scintimammography was performed in prone position 5-10min after intravenous injection of 740MBq of technetium-99m-methoxy-isobtyl-isonitrile (99mTc-MIBI) with acquisition time of 10min for every lateral and anterior projection. Digital mammography was done in all women according to a standard clinical protocol. Final diagnosis was established by histopathology. Multicentric breast cancer was defined as 2 or more distinct invasive tumors located in more than one quadrant or additional lesions from the primary tumor of more than 4cm in size. RESULTS: According to histopathological examinations multicentric breast cancer was diagnosed in 47 of 367 (12.8%) patients. Scintimammography was more effective than mammography in detecting multicentric breast cancer: sensitivity: 83.0% vs 40.4% (P<0.001), specificity: 97.8% vs 95.3% (P=0.4), accuracy: 95.9% vs 88.3% (P<0.001), positive and negative predictive values: 84.8% vs 55.9% (P=0.004) and 97.5% vs 91.6% (P<0.001), respectively. Combination of DM and SMG was characterized by increased sensitivity (93.6%), worse specificity (93.4%), accuracy (93.4%) and worse predictive value (67.7%) as compared to only SMG. CONCLUSION: Scintimammography was significantly much better by all statistical parameters than DM in the detection of multicentric breast cancer. High positive predictive value of scintimammography (84.8%) advocates it as a tool for surgery and radiotherapy planning.

Intratumoral Heterogeneity of SMAD4 Immunohistochemical Expression and Its Role in Prediction of Recurrence Pattern in Patients with Resectable Pancreatic Cancer.

BACKGROUND: The aim of our study was to evaluate consistency of SMAD4 expression in different tumor areas and its correlation with recurrence pattern in patients after resection for pancreatic cancer (PC). METHODS: Records of patients who underwent resection for nonmetastatic PC between 2001 and 2015 were analyzed. Formalin-fixed, paraffin-embedded tissue sections from different areas of primary tumor and lymph node metastases were analyzed immunohistochemically (IHC) for SMAD4 expression using TMA technology. RESULTS: SMAD4 expression was assessed in 356 tissue sections obtained from 91 patients. SMAD4 expression was positive in all assessed tumor slides only in 7 of 26 patients (26.9%). There were 54 recurrences (9 locoregional, 41 distant, and 4 both local and distant) with median follow-up of 21.7 months. There was no correlation between SMAD4 expression and locoregional recurrence pattern (p = 0.30). SMAD4 status influenced neither distant recurrence-free survival (p = 0.99) nor overall survival (p = 0.13). CONCLUSIONS: Different areas inside primary tumor and lymph node metastases express SMAD4 heterogeneously. SMAD4 IHC expression is not a biomarker of the recurrence pattern after surgical resection for PC.

Genotypic Variation of Glucosinolates and Their Breakdown Products in Leaves of Brassica rapa.

An in-depth glucosinolate (GLS) profiling was performed on a core collection of 91 Brassica rapa accessions, representing diverse morphotypes of heterogeneous geographical origin, to better understand the natural variation in GLS accumulation and GLS breakdown product formation. Leaves of the 91 B. rapa accessions were analyzed for their GLS composition by UHPLC-DAD and the corresponding breakdown products by GC-MS. Fifteen different GLSs were identified, and aliphatic GLSs prevailed regarding diversity and concentration. Twenty-three GLS breakdown products were identified, among them nine isothiocyanates, ten nitriles, and four epithionitriles. Epithionitriles were the prevailing breakdown products due to the high abundance of alkenyl GLSs. The large scale data set allowed the identification of correlations in abundance of specific GLSs or of GLS breakdown products. Discriminant function analysis identified subspecies with high levels of similarity in the acquired metabolite profiles. In general, the five main subspecies grouped significantly in terms of their GLS profiles.

The high frequency of HIV type 1-specific cellular immune responses in seronegative individuals with parenteral and/or heterosexual HIV type 1 exposure.

Some individuals remain HIV seronegative despite repeated unprotected exposure to the virus. Recent observations led to a concept that acquired immunity plays a role in protection or at least in altered susceptibility to HIV-1 infection in highly exposed seronegative (ESN) individuals. Our aim was to study HIV-specific cellular immune responses induced in parenterally and/or heterosexually ESN individuals. Nine seronegative injection drug users (IDUs), 10 seronegative individuals, and nine of their HIV-positive sexual and/or IDU partners from the cohort of IDUs were included in the study. The discordant couples had unprotected sex, and some of seronegative partners also had parenteral exposure. Cell-mediated responses were measured in peripheral blood mononuclear cells (PBMCs) by ex vivo interferon (IFN)-γ-ELISpot and ICS combining IFN-γ, tumor necrosis factor (TNF)-α, and interleukin (IL)-2 after stimulation with four consensus peptide pools (Nef, Gag, RT, Env, subtype A-EE). Thirteen out of 19 (68%) seronegative study subjects had strong Nef peptide pool-specific ELISpot responses, three (16%) subjects responded against the Gag peptide pool, and one subject had an RT peptide pool response. Nef peptide pool responses in ESN were as high as in seropositive subjects. The multiple HIV-specific cytokine production in both CD4(+) and CD8(+) T cells was shown for several ESN subjects. The functional profiles of the immune responses were different between seronegative and HIV-positive study groups. Whether the observed cellular responses have any protective role against HIV needs to be further investigated.

Design of a Brassica rapa core collection for association mapping studies.

A Brassica rapa collection of 239 accessions, based on two core collections representing different morphotypes from different geographical origins, is presented and its use for association mapping is illustrated for flowering time. We analyzed phenotypic variation of leaf and seed pod traits, plant architecture, and flowering time using data collected from three field experiments and evaluated the genetic diversity with a set of SSR markers. The Wageningen University and Research Centre (WUR) and the Vavilov Research Institute of Plant Industry (VIR) core collections had similar representations of most morphotypes, as illustrated by the phenotypic and genetic variation within these groups. The analysis of population structure revealed five subgroups in the collection, whereas previous studies of the WUR core collection indicated four subgroups; the fifth group identified consisted mainly of oil accessions from the VIR core collection, winter oils from Pakistan, and a number of other types. A very small group of summer oils is described, that is not related to other oil accessions. A candidate gene approach was chosen for association mapping of flowering time with a BrFLC1 biallelic CAPS marker and a BrFLC2 multiallelic SSR marker. The two markers were significantly associated with flowering time, but their effects were confined to certain morphotypes and (or) alleles. Based on these results, we discuss the optimal design for an association mapping population and the need to fix the heterogeneous accessions to facilitate phenotyping and genotyping.