Breast Cancer Molecular Subtyping in Practice: A Real-World Study of the APIS Breast Cancer Subtyping Assay in a Consecutive Series of Breast Core Biopsies.

The APIS Breast Cancer Subtyping Kit is an mRNA-based assessment of the seven parameters including three biomarkers routinely assessed in all the newly diagnosed breast cancers (BC), oestrogen receptor (ER), progesterone receptor (PR) and HER-2 and an additional four genes that create a novel proliferation signature, MKI67, PCNA, CCNA2 and KIF23. Taken together, the data are used to produce a molecular subtype for every sample. The kit was evaluated against the current standard protocol of immunohistochemistry (IHC) and/or in situ hybridisation (ISH) in breast cancer patients. The data were presented at the weekly breast multidisciplinary team (MDT) meeting. A total of 98 consecutive cases of pre-operative breast cancer core biopsies and two core biopsies of nodal metastases yielding 100 cases were assessed. IHC and APIS results were available for 100 and 99 cases. ER was concordant in 97% cases, PR was concordant in 89% and HER-2 results were concordant with IHC/ISH in 100% of the cases. Ki-67 IHC was discordant in 3% of cases when compared with MK167 alone but discordant in 24% when compared with the four-gene proliferation signature. In conclusion, our study indicates that the APIS Breast Cancer Subtyping Kit is highly concordant when compared to the results produced for ER/PR/HER-2 by IHC and/or ISH. The assay could play a role in the routine assessment of newly diagnosed breast cancer (BC) specimens.

Selenium nanoparticles conferred drought tolerance in tomato plants by altering the transcription pattern of microRNA-172 (miR-172), bZIP, and CRTISO genes, upregulating the antioxidant system, and stimulating secondary metabolism.

Drought stress is one of the major limiting factors for the production of tomato in Iran. In this study, the efficiency of selenate and Se nanoparticle (SeNP) foliar application on tomato plants was assessed to vestigate mitigating the risk associated with water-deficit conditions. Tomato plants were treated with SeNPs at the concentrations of 0 and 4 mg L-1; after the third sprays, the plants were exposed to water-deficit conditions. The foliar spraying with SeNPs not only improved growth, yield, and developmental switch to the flowering phase but also noticeably mitigated the detrimental risk associated with the water-deficit conditions. Gene expression experiments showed a slight increase in expression of microRNA-172 (miR-172) in the SeNP-treated plants in normal irrigation, whereas miR-172 displayed a downregulation trend in response to drought stress. The bZIP transcription factor and CRTISO genes were upregulated following the SeNP and drought treatments. Drought stress significantly increased the H2O2 accumulation that is mitigated with SeNPs. The foliar spraying with Se or SeNPs shared a similar trend to alleviate the negative effect of drought stress on the membrane integrity. The applied supplements also conferred drought tolerance through noticeable improvements in the non-enzymatic (ascorbate and glutathione) and enzymatic (catalase and peroxidase) antioxidants. The SeNP-mediated improvement in drought stress tolerance correlated significantly with increases in the activity of phenylalanine ammonia-lyase, proline, non-protein thiols, and flavonoid concentrations. SeNPs also improved the fruit quality regarding K, Mg, Fe, and Se concentrations. It was concluded that foliar spraying with SeNPs could mitigate the detrimental risk associated with the water-deficit conditions.

Zingiber zerumbet (L.) Roscoe ex Sm.: biotechnological advancements and perspectives.

Shampoo ginger (Zingiber zerumbet) is a multipurpose ginger that has confirmed their role as food, medicine, and for decorative purposes. The rhizome possesses zerumbone, curcuminoids, and other bioactive molecules that play crucial roles in treating several human diseases. To date, several reports are existing on the in vitro biotechnology of Z. zerumbet. The present review highlights the consolidated clarification and comprehensive explanation of in vitro biotechnological implications based on plant tissue culture for the improvement of Z. zerumbet. Studies on biotechnological involvement in shampoo ginger were primarily emphasized in the study of the last 3 decades, for instance, in vitro regeneration, micro-rhizome production, callus culture, somatic embryogenesis, ex vitro establishment, and molecular assessment of in vitro-raised clones. Moreover, this review provides insights into different in vitro culture systems and endophytes involvement in the production of secondary metabolites. This review will assist for advanced research areas related to in vitro manipulation of shampoo ginger, especially for the commercial cultivation of secondary metabolites rich clones of Z. zerumbet. Moreover, it will provide an insight into crop upgrading and breeding programs of this underutilized, aromatic, and medicinal plant for amended yield and quality. KEY POINTS: • Z. zerumbet is an aromatic spice and an ornamental • This review comprehensively assesses Z. zerumbet tissue culture • Key shortcomings and future directions of Z. zerumbet biotechnology.

Nasopharyngeal myiasis due to Cephalopina titillator in Southeastern Iran: a prevalence, histopathological, and molecular assessment.

The Cephalopina titillator is one of the most important causative agents of nasal myiasis in camels. This study aimed to explore the prevalence, histopathological effects, and molecular identification of C. titillator infestation in camels of Kerman province, South-Eastern Iran, between 2019 and 2021. The larvae were placed in 10% formalin for histopathological evaluation and species identification. Pieces of larval abdominal segments of C. titillator were selected for extraction of DNA. Partial mitochondrial CO1 genes were sequenced for final analysis. Out of the 870 camels examined, 339 (38.9%) were infested with larval stages of C. titillator. There was a significant difference between age and infection rate (P = 0.001), while no association between males and females (P = 0.074) was found. The infection rate was significantly higher in the winter (P < 0.001) than in the other seasons. In this study, different lesions depending on duration, locations, and the depth of larval adhesion notably degeneration changes, necrosis, and ulceration were observed. Also, in chronic cases, granulation tissue reactions were organized. Cephalopina titillator was confirmed by PCR sequencing analysis using mitochondrial CO1 region. A 582 bp nucleotide sequence was deposited in GenBank under the MW136151 accession number. Phylogenetic analysis of CO1 produced a single uniform sister clade to MZ209004 and MW167083 records from China and Iraq, respectively. The high prevalence of C. titillator in camels in this region and other areas of Iran declares that the country is in an endemic status and displays the existence of the potential risk for camels.

Molecular and Morphological Assessment of Septoria Species Associated with Ornamental Plants in Yunnan Province, China.

The Karst landform is the main geographic characteristic in South China. Such areas are rich in vegetation and especially suitable for growth of shrubs and herbaceous plants. In this study, 11 Septoria strains were obtained from different plants' leaves collected in the Kunming Botanical Garden, Yunnan Province, China. Based on single-gene and multi-gene analyses of five gene loci (tef1, rpb2, tub2, ITS, and LSU) and four gene regions (without LSU), these strains were found to belong to three independent phylogenetic lineages representing five species, including four novel taxa, and one new record for China. Five single gene trees were also provided to evaluate the effectiveness of each gene for discriminating the species, as a result of which tub2 was found to have the most suitable DNA barcode for rapid identification. Morphological descriptions, illustrations, and comparisons are provided for a more comprehensive assessment. Genealogical Concordance Phylogenetic Species Recognition (GCPSR) with a pairwise homoplasy index (PHI) test was used to evaluate the conclusions of the phylogenetic analyses.

Towards retrieving the Promethean treasure: a first molecular assessment of the freshwater fish diversity of Georgia.

In this study, we provide a first estimation of the molecular diversity of the freshwater fishes of Georgia. In addition to field collections, we integrated DNA barcode data obtained from recent works and public databases (BOLD and NCBI GenBank). Currently, the DNA barcode reference library for freshwater fishes of Georgia comprises 352 DNA barcodes for 50 species, 36 genera and 15 families (52% of total Georgian freshwater fish diversity), from which 162 DNA barcodes belonging to 41 species were newly generated as part of this study. A total of 22 species are reported from the Caspian Sea basin and 31 from the Black Sea basin. Amongst the studied taxa, seven species were found with large interspecific divergences (> 2%) while 11 species were found to share DNA barcodes within our dataset. In the course of the study, we found the first evidence of the existence of Gymnocephalus cernua (Linnaeus, 1758) and also confirm the second occurrence of invasive Rhinogobius lindbergi (Berg, 1933) in Georgia. Based on the evaluation of currently-available barcode data for Georgian fishes, we highlighted major gaps and research needs to further progress DNA-based biodiversity studies in Georgia. Though this study lays a solid base for DNA, based biodiversity assessment and monitoring approaches, further efforts within the recently started CaBOL (Caucasus Barcode Of Life) project are needed to obtain reference data for the species still lacking DNA barcodes.

Morpho-Molecular Assessment Indicates New Prognostic Aspects and Personalized Therapeutic Options in Sinonasal Melanoma.

Sinonasal melanoma is a rare subtype of melanoma and little is known about its molecular fingerprint. Systemic treatment options are limited, as targetable BRAF mutations are rare compared to cutaneous melanoma. Currently, metastatic sinonasal melanoma is being treated according to the guidelines of cutaneous melanoma. In this study, we investigated the molecular profile of 19 primary sinonasal melanomas, using a novel customized melanoma-specific next generation sequencing (NGS) panel (MelArray) of 190 genes. Results were correlated to histological and clinical features to further characterize this rare, aggressive type of melanoma and screen for prognostic markers and possible treatment options. Molecular profiles encompassed predominantly mutations in NRAS (25%), whereas KIT or BRAF p.V600 mutations were not detected. Tumor mutational burden was overall low. High level of copy number variations (CNVs) were associated with alterations in DNA-repair genes and shorter distant metastasis-free survival (p = 0.005). Monomorphic (vs. pleomorphic) morphology was found to be significantly associated with worse disease-specific survival (p < 0.001), however no correlation between morphology and molecular aberrations was found. A variety of alterations in different pathways were detected, justifying molecular testing and opening potential personalized treatment options in current study or compassionate use settings.

Molecular Assessment of Anaplasma marginale in Bovine and Rhipicephalus (Boophilus) microplus Tick of Endemic Tribal Belt of Coastal South Gujarat, India.

BACKGROUND AND METHODS: This study described the detection, prevalence and phylogeny of Anaplasma marginale in the bovine (cattle and buffaloes) and Rhipicephalus (Boophilus) microplus tick belonged to the tribal area of coastal South Gujarat, India, by amplifying 576 bp of major surface protein (msp) 5 gene using custom designed primers in the polymerase chain reaction (PCR). RESULTS: The PCR detection limit was up to 20 parasites/µl of blood in sensitivity experiment, and observed 100% specificity against Trypanosoma evansi, Babesia bigemina and Theileria annulata. Prevalence rate of the A. marginale in the bovine (n = 211)) was 18.48% and 6.64% (p < 0.05) as per the PCR and Giemsa stained blood smear, respectively. Febrile animals (35%) observed significantly (p < 0.05) higher incidence rate than the non-febrile (14.62%). The amplified msp5 had single cut site for the EcoR1 enzyme, upon digestion yielded two fragments of 365 and 211 bp on 1.0% agarose gel. The current sequence (KC811329) showed 100% homology and 1064 total score with the published nucleotide sequences of msp5 of A. marginale in the NCBI-BLAST study. Monophyletic relationship was observed with high bootstrap proportion (> 76% in Neighbor-Joining/ Maximum Likelihood) between the current and published nucleotide sequences in the phylogeny. Twenty out of 39 A. marginale infected bovine recorded R. (B.) microplus on their body surface, out of which 18 had detected the infection. The rickettsia was in 55%, 65% and 25% of anterior half, posterior half and egg of tick, respectively. CONCLUSION: The test detected A. marginale in a carrier, pre-symptomatic and symptomatic vertebrate hosts (cattle and buffalo) and different body parts of the starved R. (B.) microplus including its egg. The current genotype could be an explanation for the frequent outbreaks of bovine anaplasmosis in the targeted areas.

Circulating tumor DNA analysis in breast cancer: Is it ready for prime-time?

Precision Medicine is becoming the new paradigm in healthcare as it enables better resources allocation, treatment optimization with a potential side-effects reduction and consequent impact on quality of life and survival. This revolution is being catalyzed by liquid biopsy technologies, which provide prognostic and predictive information for advanced cancer patients, without the analytical and procedural drawbacks of tissue-biopsy. In particular, circulating tumor DNA (ctDNA) is gaining momentum as a clinically feasible option capable to capture both spatial and temporal tumor heterogeneity. Several techniques are currently available for ctDNA extraction and analysis, each with its preferential case scenarios and preanalytical implications which must be taken into consideration to effectively support clinical decision-making and to better highlight its clinical utility. Aim of this review is to summarize both analytical developments and clinical evidences to offer a comprehensive update on the deployment of ctDNA in breast cancer's (BC) characterization and treatment.

Multiple Biomarker Testing Tissue Consumption and Completion Rates With Single-gene Tests and Investigational Use of Oncomine Dx Target Test for Advanced Non-Small-cell Lung Cancer: A Single-center Analysis.

INTRODUCTION: First-line targeted therapies have been developed for advanced non-small-cell lung cancer (NSCLC). However, small biopsy samples pose a challenge to testing all relevant biomarkers. The present study characterized clinician-ordered single-gene lung cancer testing and evaluated tissue stewardship and the ability to successfully determine mutation status with single-gene testing or investigational use of the Oncomine Dx Target Test. MATERIALS AND METHODS: Clinician-submitted orders for 3659 single-gene tests (EGFR, ALK, ROS1, BRAF, KRAS, ERBB2, MET, RET, FGFR1) across 1402 samples at a large US-based commercial reference laboratory and 169 investigational Oncomine Dx Target Tests were retrospectively evaluated. The testing success rates and tissue consumption were evaluated by sample type, test type, and number of single-gene tests per sample. RESULTS: The large majority of lung tissue samples submitted for clinical testing were small (70.5% core needle biopsies; 10.0% fine needle aspirations). With single-gene testing, mutation status was successfully reported for ≥ 1 biomarker for 88.4% of the clinical samples. The success rates decreased and tissue consumption increased with testing of additional biomarkers. Investigational Oncomine Dx Target Tests were permitted 1 tissue slide each and demonstrated success rates similar to single-gene testing for ≥ 5 biomarkers on core needle biopsies, ≥ 4 biomarkers on fine needle aspirations, and ≥ 2 biomarkers on surgical resection specimens. CONCLUSION: Tissue stewardship is important to enable successful completion of genetic testing and informed NSCLC treatment decisions. Preliminary assessment of the investigational Oncomine Dx Target Test suggests it could facilitate access to multiple biomarker testing using small tissue samples to support therapy decisions for patients with advanced NSCLC.

Parkinson's disease and Toxoplasma gondii infection: Sero-molecular assess the possible link among patients.

We investigated the possible association between Parkinson's disease (PD), the second most common neurodegenerative disorder and Toxoplasma gondii infection, the most common neurotropic protozoan parasitic infection, using serological and molecular techniques. One hundred and fifteen patients with confirmed PD and 115 healthy subjects in the same age and sex distribution were enrolled in this study. Blood samples were taken from each participant and the sera was screened for anti-Toxoplasma antibodies (IgG and IgM). PCR assay was performed in duplicate using the primer pair targeting the B1 gene of Toxoplasma. Amplicons were directly sequenced to conduct the phylogenetic analysis. The prevalence of Toxoplasma infection based on IgG titer was 53% in case and 55.6% in the control groups, revealing no statistically significant association between Toxoplasma seropositivity and PD (OR=0.90; 95% CI=0.54-1.51; P=0.691). According to PCR assay, the prevalence of Toxoplasma infections was 19.3% in the case and 10.4% in control groups which the difference was statistically significant (OR=3.02; 95% CI=1.46-6.27; P=0.002). Multiple sequence alignment of Toxoplasma gondii isolates manifested a common haplotype by the identity: 93.6-100% and divergence: 0-6.7%. We concluded that T. gondii infection not only could not be a risk factor to PD, but even it could be concluded that patients with PD are in more risk to acquisition of infection. These results provide fresh insights into the ambiguous association between T. gondii infection and PD.

Molecular assessment of the bacterial community associated with Cassava (Manihot esculenta Crantz) cultivation in Cameroon.

Bacterial communities play an important role in nutrient cycles and plant development. Their distribution and activity may depend on location and environmental heterogeneity. This study characterized soil bacterial communities in cassava fields of Eastern (Andom) and Southern (Bityili) Cameroon using molecular tools. In both sites, two improved varieties (TMS-96/1414; TMS-92/0326) and a local variety (Local) were grown in a randomized block design. Composite bulk soils were collected at 10months after planting from cassava plots. The 16S rDNA region was amplified, MiSeq was performed and sequence data analyzed. The same 17 bacterial phyla were present in both Andom and Bityili, while Chlorobi and Deinococcus-Thermus were only specific to Andom. The phyla Proteobacteria, Planctomycetes, Actinobacteria and Acidobacteria were dominant. Although both sites shared similar phyla, the principal coordinate analysis revealed significant variations in their composition, suggesting that the functions of the bacteria in nutrients cycling are likely to differ between Andom and Bityili. Cassava yields were generally higher in Andom which also displayed a higher diversity of bacterial communities. This study provides useful information on the composition of bacterial communities in cassava fields in two agro-ecologies of Cameroon. It constitutes to our knowledge the first report describing soil bacterial communities in association with cassava growth in the country, using molecular tools.

Genomic analyses of early peri-implant bone healing in humans: a systematic review.

OBJECTIVE: The objective of the study was to systematically review the literature for studies reporting gene expression analyses (GEA) of the biological processes involved in early human peri-implant bone healing. METHODS: Electronic databases (MEDLINE, EMBASE) were searched in duplicate. Controlled and uncontrolled studies reporting GEA of human peri-implant tissues - including ≥5 patients and ≥2 time points - during the first 4 weeks of healing were eligible for inclusion. Methodological quality and risk of bias were also assessed. RESULTS: Four exploratory studies were included in reporting GEA of either tissues attached to SLA or SLActive implants after 4 to 14 days or cells attached to TiOBlast or Osseospeed implants after 3 to 7 days. A total of 111 implants from 43 patients were analyzed using validated array methods; however, considerable heterogeneity and risk of bias were detected. A consistent overall pattern of gene expression was observed; genes representing an immuno-inflammatory response were overexpressed at days 3 to 4, followed by genes representing osteogenic processes at day 7. Genes representing bone remodeling, angiogenesis, and neurogenesis were expressed concomitantly with osteogenesis. Several regulators of these processes, such as cytokines, growth factors, transcription factors, and signaling pathways, were identified. Implant surface properties seemed to influence the healing processes at various stages via differential gene expression. CONCLUSION: Limited evidence from gene expression studies in humans indicates that osteogenic processes commence within the first post-operative week and they appear influenced at various stages by implant surface properties.