Does the Mineral Composition of Volcanic Ashes Have a Beneficial or Detrimental Impact on the Soils and Cultivated Crops of Ecuador?

Agriculture is an important economic sector for Ecuador, sustained by food crops like maize, potatoes, and vegetables cultivated in the highlands while cash crops such as coffee, bananas, cacao, and palm oil are grown on the coastal plains. But, Ecuador is also a country under the influence of several natural hazards due to its geographical location, atmospheric dynamics, and geological characteristics. One of the main risks to food security is the presence of a large number of active volcanoes scattered all over the country with the most representative enemy, the falling volcanic ash. The bibliography in general highlights the potential toxicity of volcanic ash from a human health perspective, but it also negatively influences plant development at the seed's germination, as well as low crop pollination, damaged fruits, reduced leaf respiration depending on the type of crop, the developmental stage, the ash layer, and the climate. The mineral composition of the volcanic ash can also be beneficial for the soil by increasing fertility but at the same time with contrasting effects on plants due to the influence on soil characteristics such as pH, soil aeration, and biodiversity, which can detrimentally affect some crops.

The Fungal, Nutritional, and Metabolomic Diagnostics of the Oil Palm Elaeis guineensis Affected by Bud Rot Disease in Esmeraldas, Ecuador.

The oil palm Elaeis guineensis represents one of the most important crops in Ecuador. Considering that bud rot disease is deadly in Ecuador, more attention has been given to identifying possible causes for palm debility from this disease. We studied the involvement of fungi and nutrients in triggering bud rot disease in E. guineensis. Special emphasis was given to the molecules synthesized by the plant to protect against this devastating disease. Techniques like Diagnosis and Recommendation Integrated System (DRIS) and metagenomic analysis were used to understand the possible implications of biotic and abiotic factors in the development of bud rot disease in oil palm in Ecuador. Liquid chromatography-mass spectrometry (LC-MS) analysis was used to identify the phenolic protection barrier of the palm facing the disease. Our results indicate that fungi from Ascomyceta phylum were found in the tested samples. The species directly involved are different in soil compared with plants. The results indicate a deficiency of chemical elements, such as Ca, Mn, Mg, and Fe, which are responsible for palm debility from bud rot disease. More than 30 compounds with protective roles were identified in the leaves of symptomatic plants from the first stage of the infection.

Comparative Assessment of Antioxidant Activity and Functional Components of Chionanthus virginicus and Chionanthus pubescens from the Andean Region of Ecuador.

The present study aims to provide information about the antioxidant capacity and secondary metabolites from different plant parts of two species that are grown in Ecuador: Chionanthus pubescens (the Ecuadorian national tree), and Chionanthus virginicus (the fringe tree-endemic to the United States of America and adapted to Ecuador's physiographical and ecological conditions). These two species have still not been investigated for these characteristics. A comparative estimation of the antioxidant activities between the leaf, fruit, and inflorescence extracts was performed. In the quest for new medicines, the extracts were analyzed for phenolic, anthocyanin, and flavonoid content. A slight difference was observed between C. pubescens and C. virginicus flowers, the highest antioxidant activity being found in the C. pubescens leaf (DPPH IC50 = 62.8866 mg/mL, ABTS IC50 = 55.852 mg/mL, and FRAP IC50 = 2.8466 g/mL). Our results showed correlations between antioxidant activity, total phenolic content, and flavonoids. This study confirmed that the C. pubescens leaves and fruits from the Andean region of Ecuador represent a good source of antioxidants, especially due to the presence of a high content of phenolic compounds (homovanillic acid, 3,4 dimethoxyphenylacetic acid, vanillic acid, gallic acid, etc.) as determined by the HPLC-DAD method.

Risk factors in non­surviving patients with infection with carbapenemase­producing Enterobacterales strains in an intensive care unit.

Carbapenemase-producing Enterobacterales (CPE) are Gram-negative bacteria that belong to the Enterobacterales family and produce enzymes known as carbapenemases, which inhibit carbapenems, cephalosporins and penicillins. Carbapenem-resistant Enterobacterales (CRE) are resistant to carbapenems, cephalosporins and penicillins via mechanisms that may or may not produce carbapenemases. The identification of carbapenems is critical for the initiation of proper antibiotic therapy. The present case-control, retrospective study included 64 patients with CPE strains admitted to an intensive care unit between September, 2017 and October, 2021; of these, 34 patients with CPE succumbed and 30 control patients with CPE strains survived. CPE strains in the deceased patients were caused by Klebsiella spp. in 31 cases (91.2%) and Escherichia coli in 3 cases (8.8%). The univariate analysis revealed that the predictive factors associated with mortality in patients with CPE were admission with coronavirus disease 2019 (COVID-19) (P=0.001), invasive mechanical ventilation (P=0.001), and treatment with corticosteroids (P=0.006). The multivariate analysis revealed that admission with COVID-19 [odds ratio (OR), 16.26; 95% confidence interval (CI), 3.56-74.14; P≤0.05] and invasive mechanical ventilation (OR, 14.98; 95% CI, 1.35-166.22; P≤0.05) were associated with mortality as independent risk factors. Admission with COVID-19 increased the risk of mortality 16.26-fold and invasive mechanical ventilation increased the risk of mortality by 14.98-fold. On the whole, the present study demonstrates that the length of hospital duration in patients who acquired CPE did not influence mortality, whereas infection with COVID-19 increased and invasive mechanical ventilation were associated with an increased risk of mortality.

The Panoramic View of Ecuadorian Soil Nutrients (Deficit/Toxicity) from Different Climatic Regions and Their Possible Influence on the Metabolism of Important Crops.

Soil nutrients influence all stages (reproduction, growth, and development) of a plant species' life, and it is known that the deficit and/or toxicity of one or more nutrients has negative effects on the production of crops of commercial interest. Ecuador represents one of the "mega-diverse" countries in the world, with an agricultural sector of great importance, due to its contribution to the country's economy. This review provides a panoramic view of soil nutrients from different climatic regions of Ecuador and revises the importance of knowledge about the possible influence of nutrients from the soil on the plant metabolism able to influence the crop resistance against pathogens or to enrich the biological characteristics of these crops.

Variation of the Antioxidative Defense in Elaeis guineensis Jacq. Facing Bud Rot Disease in the Coastal Area of Ecuador.

Elaeis guineensis Jacq. has gained a reputation in the food industry as an incredible crop capable of supplying the world's largest edible oil production. In Ecuador, an important oil palm-producing country, this crop is affected in a high percentage by the bud rot disease, which is responsible for palm death. The main objective of the investigation was dedicated to understanding the palm defense mechanism facing bud rot disease, translated in the induction of reactive oxygen species, activation of defensive machinery comprising enzymatic and non-enzymatic antioxidative components, secondary metabolites, carotenoids accumulation in the palm during all stages of disease infection. For this, a survey was conducted in different oil palm plantations in the Esmeraldas province, one of the most representative for its highest incidence of bud rot disease. The survey completed DPPH, FRAP, ABTS, and other spectrophotometric analyses to underline the biochemical, biological, and physiological palm response front of bud rot incidence. The palm defense strategy in each disease stage could be represented by the phenolic compound's involvement, an increment of antioxidant activity, and the high enzymatic activity of phenylalanine ammonia-lyase (PAL). The results of the investigation made understandable the palm defense strategy front of this disease, respectively, the antioxidative defense and the palm secondary compounds involved.

Influence of Soil Nutrient Toxicity and Deficiency from Three Ecuadorian Climatic Regions on the Variation of Biological, Metabolic, and Nutritional Properties of Moringa oleifera Lam.

Moringa oleifera Lam. contains numerous essential constituents found in all plant parts (leaves, pods, and seeds). From all its edible parts, the leaf represents an effective remedy with high potential for medicinal applications. Ecuador is part of the new promising cultivation areas for Moringa, and therefore our study is emphasized to determine the influence of soil nutrition, toxicity (excess), and deficiency, from three main areas of this country, correlated with its climatic characteristics, on the mineral components, bioactive compounds' synthesis, and antioxidant capacity of Moringa. Different analyses were performed in soil and especially leaf samples for phytochemical content, antioxidant activity, calcium, protein, and vitamin C determination to identify the relationship between soil nutrients, abiotic conditions, and the therapeutic potential of this species cultivated in Ecuador. The obtained values using methods such as DPPH, FRAP, and ABTS showed a high antioxidant capacity of the leaves from the Coastal Ecuadorian region, related with total phenolic compounds' content (through the Folin-Ciocalteu method) and flavonoids in samples, with results obtained under the positive influence of high soil nutrients such as Ca, Mg, Mn, and Fe. We can conclude that M. oleifera from the coastal area of Ecuador presents the right environmental and soil conditions to positively influence its mineral and phytochemical content, making it suitable for incorporation into foods and medicines to solve the nutritional and medical problems in Ecuador and worldwide.

Abiotic Factors from Different Ecuadorian Regions and Their Contribution to Antioxidant, Metabolomic and Organoleptic Quality of Theobroma cacao L. Beans, Variety "Arriba Nacional".

Fine flavor cocoa is a unique category of cocoa that produces almonds with high aromatic potential and several sensory benefits that make it different from the basic or ordinary cocoas. Ecuador is the world's leader in the production and export of fine flavor cocoa, responsible for 63% of the world's total production due to the commercialization of the Arriba Nacional variety, known to possess an intense aroma that is unique in the cocoa world market. Besides its organoleptic specificity, this variety represents a source of important bioactive compounds associated with both sensory and health properties. This study evaluates the influence of an abiotic factor, nutritional soil status, on the phytochemical composition (methylxantines and phenolic compounds), and antioxidant and sensory properties of Arriba variety cocoa beans originating from three different geographical regions of Ecuador. We used the Diagnosis and Recommendation Integrated System (DRIS), Folin-Ciocalteau, high-performance liquid chromatography (HPLC), ABTS free-radical-scavenging activity, the α, α-diphenyl-ß-picrylhydrazyl free-radical-scavenging method (DPPH), and Ferric reducing antioxidant power (FRAP) analysis to reveal a significant correlation between Mn ions and total phenolic content, a positive implication of N in methylxanthine composition and antioxidant properties, and the importance of Ca, Mg, and K ions in increasing the flavonoid and anthocyanin content of raw cocoa beans. We showed that these nutritional elements can interfere with the nutraceutical and sensory properties of cocoa beans, as Cu, Mg, and K are correlated with anthocyaninic content, while Fe, Ca, P and Zn influenced the flavonoid content. We underline that the Arriba variety is suitable not only for the production of high-quality chocolate, but also for the increasing worldwide nutraceutical market, generating qualitative and competitive products.

The Edible Gray Oyster Fungi Pleurotus ostreatus (Jacq. ex Fr.) P. Kumm a Potent Waste Consumer, a Biofriendly Species with Antioxidant Activity Depending on the Growth Substrate

Nowadays, climate change is not the only threat facing our planet. There are also other types of pollution such as waste that poisons soils and water and kills plants, harming humans and animals. Sustainability represents a key issue for the actual Global Citizen. For this reason, our article is dedicated to offering biofriendly solutions to decrease wastes, give them a positive meaning, such as a substrate for an edible oyster fungus with nutritive and biological properties usefully for humans. Three types of wastes such as coconut coir, pine sawdust, and paper waste-representative symbols of pollution in Ecuador-have been tested as suitable growing substrate for the edible fungi Pleurotus ostreatus (Jacq. ex Fr.) P. Kumm by analyzing parameters such as Biological Efficiency, Mushroom Yield, and Productive Rate. The influence of these "waste" substrates on the nutritive (protein content), biological characteristic (antioxidant activity), and the content of human-health-sustaining compounds (phenols, flavonoids) were also evaluated using the Kjeldahal, DPPH, ABTS, FRAP, and Folin-Ciocalteu methods. The results indicate that all the waste products represent desirable substrates for growing the edible fungi, with more focus on coconut coir waste (one of the principal pollution problems in Ecuador), but that also achieved the increase in the fungi's desirable characteristics. Coconut coir waste could be an environmentally friendly solution that also offers for humans additional nutritive and healthy benefits.

Assessment of mitotic activity in breast cancer: revisited in the digital pathology era.

The assessment of cell proliferation is a key morphological feature for diagnosing various pathological lesions and predicting their clinical behaviour. Visual assessment of mitotic figures in routine histological sections remains the gold-standard method to evaluate the proliferative activity and grading of cancer. Despite the apparent simplicity of such a well-established method, visual assessment of mitotic figures in breast cancer (BC) remains a challenging task with low concordance among pathologists which can lead to under or overestimation of tumour grade and hence affects management. Guideline recommendations for counting mitoses in BC have been published to standardise methodology and improve concordance; however, the results remain less satisfactory. Alternative approaches such as the use of the proliferation marker Ki67 have been recommended but these did not show better performance in terms of concordance or prognostic stratification. The advent of whole slide image technology has brought the issue of mitotic counting in BC into the light again with more challenges to develop objective criteria for identifying and scoring mitotic figures in digitalised images. Using reliable and reproducible morphological criteria can provide the highest degree of concordance among pathologists and could even benefit the further application of artificial intelligence (AI) in breast pathology, and this relies mainly on the explicit description of these figures. In this review, we highlight the morphology of mitotic figures and their mimickers, address the current caveats in counting mitoses in breast pathology and describe how to strictly apply the morphological criteria for accurate and reliable histological grade and AI models.

Defining the area of mitoses counting in invasive breast cancer using whole slide image.

Although counting mitoses is part of breast cancer grading, concordance studies showed low agreement. Refining the criteria for mitotic counting can improve concordance, particularly when using whole slide images (WSIs). This study aims to refine the methodology for optimal mitoses counting on WSI. Digital images of 595 hematoxylin and eosin stained sections were evaluated. Several morphological criteria were investigated and applied to define mitotic hotspots. Reproducibility, representativeness, time, and association with outcome were the criteria used to evaluate the best area size for mitoses counting. Three approaches for scoring mitoses on WSIs (single and multiple annotated rectangles and multiple digital high-power (×40) screen fields (HPSFs)) were evaluated. The relative increase in tumor cell density was the most significant and easiest parameter for identifying hotspots. Counting mitoses in 3 mm2 area was the most representative regarding saturation and concordance levels. Counting in area <2 mm2 resulted in a significant reduction in mitotic count (P = 0.02), whereas counting in area ≥4 mm2 was time-consuming and did not add a significant rise in overall mitotic count (P = 0.08). Using multiple HPSF, following calibration, provided the most reliable, timesaving, and practical method for mitoses counting on WSI. This study provides evidence-based methodology for defining the area and methodology of visual mitoses counting using WSI. Visual mitoses scoring on WSI can be performed reliably by adjusting the number of monitor screens.

Visual assessment of mitotic figures in breast cancer: a comparative study between light microscopy and whole slide images.

BACKGROUND AND AIMS: Visual assessment of mitotic figures in breast cancer (BC) remains a challenge. This is expected to be more pronounced in the digital pathology era. This study aims to refine the criteria of mitotic figure recognition, particularly in whole slide images (WSI). METHOD AND RESULTS: Haematoxylin and eosin (H&E)-stained BC sections (n = 506) were examined using light microscopy (LM) and WSI. A set of features for identifying mitosis in WSI and to distinguish true figures from mimickers was developed. Changes in the mitotic count between the two platforms was explored. Morphological features of mitoses were recorded separately, including absence of nuclear membrane, chromatin hairy-like projections, shape, cytoplasmic features, mitotic cell size and relationship to surrounding cells. Each mitotic phase has its own mimickers. Fifty-eight per cent of mitoses showed absent hairy-like projection in WSI; however, 89% retained their ragged nuclear border, which distinguished them from mimickers including apoptotic cells, lymphocytes and dark elongated hyperchromatic structures. Mitosis in WSI showed loss of fine details, and there was a 20% average reduction rate of mitotic counts when compared to the same area on LM. Using refined mitosis recognition criteria in WSI resulted in a twofold improvement of interobserver concordance. However, when compared to LM, 19% of cases were underscored in WSIs. CONCLUSIONS: All morphological features of mitosis should be considered to enable recognition and differentiation from their mimickers, particularly in WSI, to ensure reliable BC grading. Refining mitotic cut-offs per specific area when using WSI, based on the degree of reduction and association with outcome, is warranted.

Diagnostic concordance of phyllodes tumour of the breast.

AIMS: Phyllodes tumours (PT) are rare and distinct breast tumours, which span a morphological continuum. Classification into benign, borderline and malignant categories reflects their biology and clinical behaviour and is essential to guide management. This study aims to assess the diagnostic agreement of PT using the UK National Health Service Breast Screening Programme (NHSBSP) breast pathology external quality assurance (EQA) scheme data. METHODS AND RESULTS: Twenty-six PTs were identified in the EQA scheme, which were diagnosed by an average of 607 participants/circulation. Data on diagnostic categories were collected and representative slides were reviewed. The level of concordance between reporting pathologists was assessed. There were 14 benign, six borderline and six malignant PT. The overall rate of diagnosis agreement was 86% when analysed as benign lesions, borderline PT and malignant lesions, which decreased to 79% when diagnosed as PT (irrespective of grade) and to 63% when the diagnosis was further refined to PT categories (benign, borderline and malignant PTs). The highest agreement rate was observed in malignant PT (86%) and the lowest in borderline PT (42%). Malignant heterologous elements, stromal overgrowth and leaf-like architecture are features associated with higher concordance rates. Lower-priority features were stromal expansion, clefting and multinodularity. CONCLUSION: The concordance of PT diagnosis, as an entity, is high, but its classification into benign, borderline and malignant has variable agreement levels, with borderline tumours having the lowest concordance rate. More research to refine the diagnostic criteria for categorisation of PT is warranted to improve concordance between pathologists.

Combined total internal reflection AF spectral-imaging and Raman spectroscopy for fast assessment of surgical margins during breast cancer surgery.

The standard treatment for breast cancer is surgical removal mainly through breast-conserving surgery (BCS). We developed a new technique based on auto-fluorescence (AF) spectral imaging and Raman spectroscopy for fast intraoperative assessment of excision margins in BCS. A new wide-field AF imaging unit based on total internal reflection (TIR) was combined with a Raman spectroscopy microscope equipped with a 785 nm laser. The wavelength of the AF excitation was optimized to 365 nm in order to maximize the discrimination of adipose tissue. This approach allows for the non-adipose regions of tissue, which are at a higher risk of containing a tumor, to be targeted more efficiently by the Raman spectroscopy measurements. The integrated TIR-AF-Raman was tested on small tissue samples as well as fresh wide local excisions, delivering the analysis of the entire cruciate surface of BCS specimens (5.1 × 7.6 cm2) in less than 45 minutes and also providing information regarding the location of the tumor in the specimen. Full automation of the instrument and selection of a faster translation stage would allow for the measurement of BCS specimens within an intraoperative time scale (20 minutes). This study demonstrates that the TIR-AF Raman microscope represents a feasible step towards the development of a technique for intraoperative assessment of large WLE within intraoperative timescales.

Correlations of morphological features and surgical management with clinical outcome in a multicentre study of 241 phyllodes tumours of the breast.

AIMS: Phyllodes tumours (PTs) represent an unusual but complex group of breast lesions with a tendency to recur locally and, less commonly, metastasise. On core biopsies, their appearances can be difficult to discriminate from those of other fibroepithelial lesions, which may compromise their surgical management. The aims of this study were to assess the preoperative diagnosis of PTs and to evaluate the impacts of surgical management and morphological features on their behaviour. METHODS AND RESULTS: We combined datasets from three centres over two decades, including core biopsies, excision specimens, and follow-up. Core biopsy results were compared with final excision specimens. The relationships of surgical procedure and morphological features with local recurrence (LR) and metastasis were assessed. Two hundred and forty-one PTs were studied. Core biopsy resulted in a diagnosis of possible or definite PT in 76% of cases. Malignant tumours were more likely to be larger, occurred at an older age, and were surgically more challenging, with difficulties being encountered in achieving negative margins. There were 12 cases (5%) that showed LR alone, and another six cases (2.5%) that had distant metastases. Morphological features associated with adverse outcome were grade of PT, increased mitotic counts, necrosis, infiltrative margins, stromal atypia, and heterologous components. Both LR and metastatic behaviour correlated with larger size and distance to margins. CONCLUSIONS: Our results suggest that excision margins have a significant impact on LR of PT, whereas metastatic behaviour is influenced by tumour biology. We add to the evidence base on histological features of tumours that contribute to long-term outcomes of PT patients.

Metaplastic carcinomas of the breast without evidence of epithelial differentiation: a diagnostic approach for management.

AIMS: Although rare, malignant sarcomatoid breast tumours without evidence of epithelial differentiation comprise a diagnostic challenge with management implications. Earlier studies have generally considered these to be primary breast sarcomas; however, supporting evidence is lacking and management remains variable. This study aimed to provide an evidence-based approach to improve the consistency of diagnosis and management for such cases. METHODS AND RESULTS: A large series (n = 140) of metaplastic breast carcinoma (MBC) diagnosed in Nottingham over 18 years was analysed. Only cases with available data on immunohistochemical expression of cytokeratins (CKs) were included. The prevalence and pattern of expression for various CKs were assessed and details of tumours negative for CKs were collected. A diagnostic approach based on our experience is provided. Forty-seven cases (34%) showed foci of conventional type invasive breast carcinoma or ductal carcinoma in situ (DCIS), while 93 cases (66%) were diagnosed as MBC based on morphology and/or CK expression. Ninety-seven cases (69%) were negative for one or more CKs, with 18 cases (13%) negative for five or more CKs. Eight cases (6%) lacked expression of all CKs tested. Further examination showed evidence of carcinomatous nature in five cases, and three were diagnosed as MBC following extensive diagnostic work-up and based on our experience. CONCLUSION: This study suggests that MBC represents a spectrum of neoplasms, with some lacking CK expression. Sarcomatoid neoplasms of the breast lacking evidence of carcinomatous morphology and CK expression may represent an extreme end of differentiation that can be considered as carcinomas rather than sarcomas for management purposes (following extensive work-up).

Visual histological assessment of morphological features reflects the underlying molecular profile in invasive breast cancer: a morphomolecular study.

AIMS: Tumour genotype and phenotype are related and can predict outcome. In this study, we hypothesised that the visual assessment of breast cancer (BC) morphological features can provide valuable insight into underlying molecular profiles. METHODS AND RESULTS: The Cancer Genome Atlas (TCGA) BC cohort was used (n = 743) and morphological features, including Nottingham grade and its components and nucleolar prominence, were assessed utilising whole-slide images (WSIs). Two independent scores were assigned, and discordant cases were utilised to represent cases with intermediate morphological features. Differentially expressed genes (DEGs) were identified for each feature, compared among concordant/discordant cases and tested for specific pathways. Concordant grading was observed in 467 of 743 (63%) of cases. Among concordant case groups, eight common DEGs (UGT8, DDC, RGR, RLBP1, SPRR1B, CXorf49B, PSAPL1 and SPRR2G) were associated with overall tumour grade and its components. These genes are related mainly to cellular proliferation, differentiation and metabolism. The number of DEGs in cases with discordant grading was larger than those identified in concordant cases. The largest number of DEGs was observed in discordant grade 1:3 cases (n = 1185). DEGs were identified for each discordant component. Some DEGs were uniquely associated with well-defined specific morphological features, whereas expression/co-expression of other genes was identified across multiple features and underlined intermediate morphological features. CONCLUSION: Morphological features are probably related to distinct underlying molecular profiles that drive both morphology and behaviour. This study provides further evidence to support the use of image-based analysis of WSIs, including artificial intelligence algorithms, to predict tumour molecular profiles and outcome.

Immunohistochemical assessment of HRAS Q61R mutations in breast adenomyoepitheliomas.

AIMS: Breast adenomyoepitheliomas (AMEs) are uncommon tumours. Most oestrogen receptor (ER)-positive AMEs have mutations in phosphoinositide 3-kinase (PI3K) pathway genes, whereas ER-negative AMEs usually harbour concurrent mutations affecting the HRAS Q61 hotspot and PI3K pathway genes. Here, we sought to determine the sensitivity and specificity of RAS Q61R immunohistochemical (IHC) analysis for detection of HRAS Q61R mutations in AMEs. METHODS AND RESULTS: Twenty-six AMEs (14 ER-positive; 12 ER-negative) previously subjected to massively parallel sequencing (n = 21) or Sanger sequencing (n = 5) of the HRAS Q61 hotspot locus were included in this study. All AMEs were subjected to IHC analysis with a monoclonal (SP174) RAS Q61R-specific antibody, in addition to detailed histopathological analysis. Nine ER-negative AMEs harboured HRAS mutations, including Q61R (n = 7) and Q61K (n = 2) mutations. Five of seven (71%) AMEs with HRAS Q61R mutations were immunohistochemically positive, whereas none of the AMEs lacking HRAS Q61R mutations (n = 17) were immunoreactive. RAS Q61R immunoreactivity was restricted to the myoepithelium in 80% (4/5) of cases, whereas one case showed immunoreactivity in both the epithelial component and the myoepithelial component. RAS Q61R immunohistochemically positive AMEs were associated with infiltrative borders (P < 0.001), necrosis (P < 0.01) and mitotic index in the epithelial (P < 0.05) and myoepithelial (P < 0.01) components. RAS Q61R IHC assessment did not reveal Q61K mutations (0/2). CONCLUSIONS: IHC analysis of RAS Q61R shows high specificity (100%) and moderate sensitivity (71%) for detection of HRAS Q61R mutations in breast AMEs, and appears not to detect HRAS Q61K mutations. IHC analysis of RAS Q61R may constitute a useful technique in the diagnostic workup of ER-negative AMEs.