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Anabolic-androgenic steroids (AAS) abuse is linked to some abnormalities in several tissues including the kidney. However, the precise molecular mediators involved in AAS-induced kidney disorder remain elusive. The main objective of the present study was to investigate the effect of Nandrolone decanoate on kidney injury alone or in combination with moderate exercise and its related mechanisms. Thirty-two male Wistar rats were subdivided randomly into four groups. control (Con), Nandrolone (10 mg/kg)(N), Exercise (Exe), Nandrolone + Exercise (N+Exe). RESULTS: After 6 weeks, nandrolone treatment led to a significant increase in functional parameters such as serum cystatin c, urea, creatinine, albuminuria and Albumin/ creatinine ratio indicating kidney dysfunction. Moreover, nandrolone treatment increased vacuolization, focal inflammation, hemorragia, cast formation fibrosis in the renal tissue of rats. miRNA-146a increased in kidney tissue after nandrolone exposure by using RT-PCR which may be considered idealtheranomiRNAcandidates for diagnosis and treatment. Western blotting indicated that IRAK1, TRAF6, TNF-α, NF-κB, iNOS and TGF-ß protein expressions were considerably elevated in the kidneys of nandrolone treated rats. Moderate exercise could alleviate the renal dysfunction, histological abnormalities and aforementioned proteins. Our findings suggested that nandrolone consumption can cause damage to kidney tissue probably through miRNA-146a targeting IRAK1 and TRAF6 via activation of the NF-κB and TGF-ß pathway. These results provide future lines of research in the identification of theranoMiRNAs related to nandrolone treatment, which can be ameliorated by moderate exercise.

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Α sensitive and selective derivatization and inject method for the quantification of intact nandrolone phase II oxo-metabolites was developed and validated using liquid chromatography - (tandem high resolution) mass spectrometry (LC-MS/(HRMS)). For the derivatization, Girard's reagent T (GRT) was used directly in natural urine samples and the analysis of the metabolites of interest was performed by direct injection into LC-MS/(HRMS) system operating in positive ionization mode. Derivatization enabled the efficient detection of nandrolone oxo-metabolites, while at the same time producing intense product ions under collision-induced dissociation (CID) conditions that are related to metabolites of the steroid backbone and not to the conjugated moieties. Glucuronide and sulfate metabolites of nandrolone were chromatographically resolved and quantified in the same run in the range of 1-100 ng mL-1, while at the same time structure identification could be performed for each metabolite. Full validation of the method was performed according to the World Anti-Doping Agency (WADA) International Standard for Laboratories (ISL). Nandrolone oxo-metabolites were quantified in two sets of urine samples, the first set consisted of real urine samples previously detected as negative and the second set consisted of urine samples collected from two excretion studies after nandrolone decanoate administration. The results for 19-norandrosterone glucuronide (19-NAG) and 19-noretiocholanolone glucuronide (19-NEG) were compared with those obtained by traditional gas chromatography - (tandem) mass spectrometry (GC-MS/[MS]) method.

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Nandrolone (NT) is a type of androgen anabolic steroid that is often illegally used in cattle farming, leading to unpredictable harm to human health via the food chain. In this study, a rapid detection method for NT in the samples of cattle farming was established using a portable mass spectrometer. The instrument parameters were optimized, including a thermal desorption temperature of 220 °C, a pump speed of 30 %, an APCI ionization voltage of 3900 v, and an injection volume of 6 µL. The samples of bovine urine, feed, sewage, and tissue were selected, and extracted using a solution of methanol:acetonitrile (1:1, v/v), followed by spiking a NT standard solution (1000 ng·mL-1) and ionization through the APCI ion source for detection. The results showed that NT could not be detected in beef and feed due to the complexity of the matrix, while clear signals of NT ions were observed in bovine urine and sewage samples, with LODs of 1000 and 100 ng·mL-1, respectively. Furthermore, quantitative analysis was attempted, and a good linear relationship (R2 = 0.9952) was observed for NT in sewage within the range of 100 to 1000 ng·mL-1. At spiked levels of 100, 500, 1000 and 2000 ng mL-1, the recovery rates ranged from 74.3 % to 92.8 %, with a relative standard deviation (n = 6) of less than 15 %. In conclusion, this detection method offers the advantages of simplicity, rapidity, strong timeliness, and specificity, making it suitable for on-site detection. It can be used for qualitative screening of nandrolone in bovine urine and quantitative analysis of nandrolone in sewage.

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Caffeine and anabolic-androgenic steroids (AAS) are commonly used to improve muscle mass and athletic performance. Nandrolone Decanoate (ND) is one of the most abused AAS worldwide, leading to behavioral changes in both humans and rodents. Caffeine, the most widely consumed psychostimulant globally, is present in various thermogenic and gym supplements. Low and moderate doses of caffeine antagonize adenosine receptors and have been linked to improved memory and pain relief. We have previously demonstrated that consuming caffeine prevents the risk-taking behavior triggered by nandrolone. In this study, we aimed to investigate the long-term effects of ND and caffeine, either alone or in combination, on passive avoidance memory and nociception. We used the step-down and hot-plate tasks in male and female Lister Hooded rats. Our results confirmed the antinociceptive effect of caffeine and indicated that chronic administration of the ND-caffeine association promotes the evocation of aversive memory in female rats.

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Boldenone (Bdn) and nandrolone (Ndn) are anabolic androgenic steroids (AASs) that, as our previous studies have shown, may increase the risk of neoplastic transformation of porcine ovarian putative stem cells (poPSCs). The NF-κB pathway may be important in the processes of carcinogenesis and tumour progression. Therefore, in this work, we decided to test the hypothesis of whether Bdn and Ndn can activate the NF-κB pathway by acting through the membrane androgen receptor ZIP-9. For this purpose, the expression profiles of both genes involved in the NF-κB pathway and the gene coding for the ZIP-9 receptor were checked. The expression and localization of proteins of this pathway in poPSCs were also examined. Additionally, the expression of the ZIP-9 receptor and the concentration of the NF-κB1 and 2 protein complex were determined. Activation of the NF-κB pathway was primarily confirmed by an increase in the relative abundances of phosphorylated forms of RelA protein and IκBα inhibitor. Reduced quantitative profiles pinpointed not only for genes representing this pathway but also for unphosphorylated proteins, and, simultaneously, decreased concentration of the NF-κB1 and 2 complex may indicate post-activation silencing by negative feedback. However, the remarkably and sustainably diminished expression levels noticed for the SLC39A9 gene and ZIP-9 protein suggest that this receptor does not play an important role in the regulation of the NF-κB pathway.

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Nandrolone and its prohormones, including 19-norandrost-4-ene-3,17-dione and 19-norandrost-4-ene-3ß,17ß-diol, are anabolic steroids forbidden at all times in sports according to the World Anti-Doping Code Prohibited List and its metabolite 19-norandrosterone (19NA) is the preferred urinary target compound to identify their abuse. In recent years, an increasing number of 19NA isotope ratio mass spectrometry (IRMS) cases have arisen that, based on the initial testing procedure, were likely to result in an adverse analytical finding but were concluded negative after IRMS analysis. The current study was therefore set up to gain a better insight on the prevalence of nandrolone preparations with endogenous carbon isotope ratio values in Australia. Suitable workplace (non-athlete) urine samples that had previously been reported positive for 19NA were identified and analysed on IRMS. A total of 82% of the samples that were analysed were reported with enriched carbon isotope ratios of 19NA (i.e., 19NA greater than -26‰). This indicates that there is a high prevalence of nandrolone-containing anabolic androgenic steroid preparations in Australia that have 'endogenous' carbon isotope ratios which reduces the ability to identify exogenous nandrolone.

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Long-term use of anabolic androgenic steroids (AAS) in supratherapeutic doses is associated with severe adverse effects, including physical, mental, and behavioral alterations. When used for recreational purposes several AAS are often combined, and in scientific studies of the physiological impact of AAS either a single compound or a cocktail of several steroids is often used. Because of this, steroid-specific effects have been difficult to define and are not fully elucidated. The present study used male Wistar rats to evaluate potential somatic and behavioral effects of three different AAS; the decanoate esters of nandrolone, testosterone, and trenbolone. The rats were exposed to 15 mg/kg of nandrolone decanoate, testosterone decanoate, or trenbolone decanoate every third day for 24 days. Body weight gain and organ weights (thymus, liver, kidney, testis, and heart) were measured together with the corticosterone plasma levels. Behavioral effects were studied in the novel object recognition-test (NOR-test) and the multivariate concentric square field-test (MCSF-test). The results conclude that nandrolone decanoate, but neither testosterone decanoate nor trenbolone decanoate, caused impaired recognition memory in the NOR-test, indicating an altered cognitive function. The behavioral profile and stress hormone level of the rats were not affected by the AAS treatments. Furthermore, the study revealed diverse AAS-induced somatic effects i.e., reduced body weight development and changes in organ weights. Of the three AAS included in the study, nandrolone decanoate was identified to cause the most prominent impact on the male rat, as it affected body weight development, the weights of multiple organs, and caused an impaired memory function.

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Osteodystrophia fibrosa is a metabolic disease of goats resulting from the insufficient mineralization of bones, excessive bone resorption, and improper bone development, leading to subsequent accumulation of the fibrous connective tissue. This report describes the successful therapeutic management of acute osteodystrophia fibrosa in early-weaned goat kids by following a medical protocol. Three Malabari male goat kids of forty days old from two different litters of the same farm were presented with spontaneous, bilaterally symmetrical, non-inflammatory facial swelling, protruded tongue, prominent eyeballs, frothy drooling saliva, diarrhoea, and inanition due to the inability in mastication or suckling for the last two weeks. As per the history, these kids were weaned and started feeding pelleted concentrate feed and jackfruit leaves along with the milk sucking three weeks back only. On clinical examination, the vital parameters were normal and the key phenotypic abnormalities noted were the abnormally developed maxilla/mandible, manifested as a swelling that was hard to touch, and the loosely attached teeth. In biochemical evaluation, the reduction in Ca:P ratio was evident and the serum creatinine level was within normal ranges. A therapeutic protocol was devised for three weeks by including calcium (calcium gluconate), phosphate-binder (Kaolin), vitamins (Vit. ADEH), and anabolic steroid (Nandrolone; a well-established drug promoting formation and mineralization of bone in mammals). Considerable improvement was observed after three weeks of therapy and the animals were having apparently normal facial appearance on the review after three months. The kids showed normal healthy growth and were slaughtered later for meat purpose after gaining adult body weight. This report accentuates the possibility of acute hypocalcemia-induced osteodystrophia fibrosa, rather than hyperphosphatemia-induced, and its management using anabolic steroids for better clinical recovery in growing goat kids.

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The aim of this study was to investigate the effects of 10 mg/kg/week of nandrolone decanoate (DECA - Deca Durabolin®) on body composition, hormonal levels, spermatic parameters, redox status, and morphometric parameters of testicle and epididymis; furthermore, the fertility capacity of Wistar rats was measured thought in vitro fertilization (IVF). The animals (n = 16) were divided into two groups: control group (CTRL, n = 8), which received only vehicle composed by peanut oil and 10% of the benzoic alcohol and nandrolone decanoate group (DECA, n = 8), which received intramuscular injections of DECA for 8 weeks, both groups were treated for 8 weeks. The results demonstrate significative decrease in visceral fat, testosterone levels, and thiol content on epididymis, reduction on normal sperm parameters, and deleterious effect on testicles and epididymis tissue morphology showing reduction of germ height and luminal diameter on the DECA group. Thus, it can be concluded that high doses of nandrolone decanoate impairs male reproductive parameters.

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Background and Objectives: Nandrolone decanoate (ND) is the most widely used among the anabolic androgenic steroids (AAS), synthetic substances derived from testosterone, to improve muscular and health gains associated with exercises. The AAS leads to physical performance enhancement and presents anti-aging properties, but its abuse is associated with several adverse effects. Supraphysiological doses of AAS with or without physical exercise can cause morphological and functional alterations in neuromuscular interactions. This study aims to investigate the effects of ND supraphysiological doses in neuromuscular interactions, focusing on the soleus muscle and its neuromuscular junctions (NMJs) in rats, associated or not with physical exercise. Materials and Methods: Forty male Sprague Dawley rats were divided into four groups: sedentary and exercised groups, with or without ND at the dose of 10 mg/kg/week. The animals were treated for eight weeks, with intramuscular injections, and the soleus muscle was collected for morphological analyses. Results: The supraphysiological doses of ND in the sedentary group caused muscle degeneration, evidenced by splitting fibers, clusters of small fibers, irregular myofibrils, altered sarcomeres, an increase in collagen deposition and in the number of type I muscle fibers (slow-twitch) and central nuclei, as well as a decrease in fibers with peripheral nuclei. On the other hand, in the ND exercise group, there was an increase in the NMJs diameter with scattering of its acetylcholine receptors, although no major morphological changes were found in the skeletal muscle. Thus, the alterations caused by ND in sedentary rats were partially reversed by physical exercise. Conclusions: The supraphysiological ND exposure in the sedentary rats promoted an increase in muscle oxidative pattern and adverse morphological alterations in skeletal muscle, resulting from damage or post-injury regeneration. In the ND-exercised rats, no major morphological changes were found. Thus, the physical exercise partially reversed the alterations caused by ND in sedentary rats.

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Nandrolone (Estr-4-en-17ß-ol-3-one) is a derivative of testosterone and a naturally occurring anabolic-androgenic agent which belongs to the steroid group. Crystal structures of four short, medium and long esterified forms of nandrolone, including propionate, phenylpropionate, cypionate and undecanoate were determined using single-crystal X-ray diffraction. Crystal packing, supramolecular features and intermolecular interactions were described based on a quantitative and qualitative Hirshfeld surfaces analysis accompanied by evaluation of crystal energies and intermolecular interactions computation. Also, the solubility of the esters was investigated from a pharmaceutical perspective.

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A 41-year-old man was admitted to the Emergency Department with bilateral numbness in lower extremities and left femoral palsy. He also reported constitutional symptoms, vomiting and non-bloody diarrhoea for the last several months. He had a previous history of eosinophilic asthma with erratic follow-up. During admission, eosinophil count was 66% of white blood cells. Sural nerve biopsy revealed vasculitis with eosinophilic infiltration. Further evaluations consisted of colonoscopy and nasal endoscopy that confirmed eosinophil infiltrates on colonic ulcers and nasal polypi. The patient was started on systemic corticosteroids and cyclophosphamide. Among his personal records, he had been taking nandrolone decanoate without medical prescription, and had withdrawn a few years before the first asthma exacerbation.

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Skeletal muscle undergoes rapid and extensive atrophy following nerve transection though the underlying mechanisms remain incompletely understood. We previously showed transiently elevated Notch 1 signaling in denervated skeletal muscle that was abrogated by administration of nandrolone (an anabolic steroid) combined with replacement doses of testosterone. Numb is an adaptor molecule present in myogenic precursors and skeletal muscle fibers that is vital for normal tissue repair after muscle injury and for skeletal muscle contractile function. It is unclear whether the increase in Notch signaling observed in denervated muscle contributes to denervation and whether expression of Numb in myofibers slows denervation atrophy. To address these questions, the degree of denervation atrophy, Notch signaling, and Numb expression was studied over time after denervation in C57B6J mice treated with nandrolone, nandrolone plus testosterone or vehicle. Nandrolone increased Numb expression and reduced Notch signaling. Neither nandrolone alone nor nandrolone plus testosterone changed the rate of denervation atrophy. We next compared rates of denervation atrophy between mice with conditional, tamoxifen-inducible knockout of Numb in myofibers and genetically identical mice treated with vehicle. Numb cKO had no effect on denervation atrophy in this model. Taken together, the data indicate that loss of Numb in myofibers does not alter the course of denervation atrophy and that upregulation of Numb and blunting of the denervation-atrophy induced activation of Notch do not change the course of denervation atrophy.

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BACKGROUND: Anabolic steroids (AS) are commonly abused by body builders and athletes aiming to increase their strength and muscle mass but unfortunately, the long-term use of AS may lead to serious side effects. Nandrolone Decanoate is one of the Class II anabolic androgenic steroids which quickly spread globally and used clinically and illicitly. Our research was directed to assess the toxic effects of anabolic steroids on cardiac and skeletal muscles in male albino rats and to evaluate the potential ameliorative effects of fenugreek seeds extract and silymarin. METHODS: Our research was done on 120 male albino rats that were allocated into 6 groups; group I: Served as a control group, group II: Received the anabolic steroid Nandrolone Decanoate, group III: Received silymarin orally, group IV: Received fenugreek seeds extract orally, group (V): Received the anabolic steroid Nandrolone Decanoate and silymarin and group (VI): Received the anabolic steroid Nandrolone Decanoate and fenugreek seeds extract. By the end of the study, rats were sacrificed, and blood samples were collected for biochemical analysis and autopsy samples for histopathological examination. RESULTS: The anabolic steroids toxic effects on rats showed a significant decrease in serum High Density Lipoprotein (HDL) level and increase in cholesterol, triglycerides, and Low-Density Lipoprotein (LDL) levels. There was a significant elevation in cardiac troponin I level. As regards to histopathological examination of the cardiac and skeletal muscles, the study showed marked degenerative changes and necrosis. Both silymarin and fenugreek seeds extract provided a protective effect on the biochemical and histopathological changes. The antioxidant effects of silymarin and fenugreek seeds extract were evaluated on the heart, skeletal muscles and showed that, the tissue levels of Superoxide dismutase (SOD), Catalase and reduced glutathione (GSH) decreased in AS treated rats compared to the control group. On the other hand, the tissue Malondialdehyde (MDA) levels were elevated. CONCLUSIONS: Anabolic steroids have a toxic effect on the cardiac and skeletal muscles of albino rats with improvement by treatment with fenugreek seeds extract and silymarin.

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Nandrolone, an anabolic androgenic steroid, is included in the prohibited list of the World Anti-Doping Agency. Drugs of abuse activate brain dopamine neurons and nandrolone has been suspected of inducing dependence. Accordingly, possible critical periods for the effects of nandrolone on muscular strength and dopaminergic activity have been investigated, including the effects of chronically administered nandrolone alone and on morphine-induced increases in dopamine efflux in the nucleus accumbens. Six- or 10-week-old male Sprague-Dawley rats were used. Treatment with nandrolone was initiated in adolescent (6-week-old) and young adult (10-week-old) rats. Nandrolone (5.0 mg/kg s.c.) or sesame oil vehicle was given once daily, on six consecutive days per week, for 3 weeks and then once per day for 4 consecutive days. Nandrolone enhanced the developmental increase in grip strength of 6- but not 10-week-old rats, without altering the developmental increase in body weight of either age group. Using in vivo microdialysis in freely moving 6-week-old rats given nandrolone for 4 weeks, basal accumbal dopamine efflux was unaltered, while the increase in dopamine efflux induced by acute administration of morphine (1.0 mg/kg s.c.) was reduced. The present study provides in vivo evidence that adolescence constitutes a critical period during which repeated administration of nandrolone enhances increases in muscular strength without influencing increases in body weight. Though repeated administration of nandrolone during this period of adolescence did not stimulate in vivo mesolimbic dopaminergic activity, it disrupted stimulation by an opioid, the drug class that is most commonly coabused with nandrolone.

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Recently, our research group have focused on an interesting project in which a novel dual template molecularly imprinted (DTMIP) biosensor was fabricated and assisted by second-order differential pulse voltammetric (DPV) data for simultaneous determination of nandrolone decanoate (ND) and testosterone decanoate (TS). An indium tin oxide (ITO) was modified with multiwalled carbon nanotubes-graphene-ionic liquid (MWCNT-Gr-IL) and then, the fullerene C60 was casted onto the surface of MWCNT-Gr-IL/ITO and electrochemically reduced. Finally, DTMIPs were electrosynthesized by electropolymerization of 4-aminobenzoic acid (ABA) as monomer with ND and TS as template molecules to obtain the final structure of the biosensor (DTMIP/C60/MWCNT-Gr-IL/ITO). Structure of the biosensor was electrochemically and microscopically characterized. The ND and TS generated two severely overlapped DPVs at the surface of the biosensor which forced us to assist the biosensor with three-way calibration by second-order DPV data to simultaneous determine them. Two second-order algorithms including multivariate curve resolution alternating least squares (MCR-ALS) and parallel factor analysis2 (PARAFAC2) were used to build second-order calibration models and evaluation of their performance in the analysis of synthetic samples showed more superiority of the MCR-ALS than PARAFC2 which motivated us to select PARAFC2 for the analysis of urine samples as real cases. Application of the biosensor assisted by PARAFC2 for the analysis of urine samples towards simultaneous determination of ND and TS was successful.

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AIMS: It has been revealed that membrane androgen receptor activation modulates avoidance memory and synaptic plasticity. In a previous study, we showed that Calcineurin, a calcium dependent phosphatase, could be a potential mediator of these AR effects. Also, it is reported that AR activation leads to L-type calcium channel activation. The aim of the current study is to test whether L-type calcium channels are downstream of AR and whether this signal pathway mediates the impairment effect of androgenic steroids on passive avoidance memory and synaptic plasticity. MATERIALS AND METHODS: We measured the effect of Nandrolone Decanoate (AR agonist), AR antagonist (Nilutamide) plus ND or L-type calcium channel inhibitor (Nifedipine) plus ND on passive avoidance performance of adolescent male rats. For extracellular field potential recordings hippocampal slices were perfused with ND, Nilutamide-ND or Nifedipine-ND. KEY FINDINGS: Our results clarified that AR activation by ND could impair avoidance behavior as step through latency decreased in ND-treated group while application of both Nilutamide and Nifedipine reestablished normal avoidance behavior. Also, LTP induction in the CA1 area of hippocampus was diminished by ND perfusion and both AR antagonist and L-type calcium channel inhibitor application lead to normal LTP. These findings support our hypothesis that activation of L-type calcium channels are involved in ARs mechanism effects on both avoidance behavior and hippocampal synaptic plasticity. SIGNIFICANCE: Understanding the biological effects of AR agonists on cognitive processes and its cellular mechanism may be a new/supplementary way to treating fear-related disorders.

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Anabolic-androgenic steroids (AAS) and caffeine can induce several behavioral alterations in humans and rodents. Administration of nandrolone decanoate is known to affect defensive responses to aversive stimuli, generally decreasing inhibitory control and increasing aggressivity but whether caffeine intake influences behavioral changes induced by AAS is unknown. The present study aimed to investigate behavioral effects of caffeine (a non-selective antagonist of adenosine receptors) alone or combined with nandrolone decanoate (one of the most commonly AAS abused) in female and male Lister Hooded rats. Our results indicated that chronic administration of nandrolone decanoate (10 mg/kg, i.m., once a week for 8 weeks) decreased risk assessment/anxiety-like behaviors (in the elevated plus maze test), regardless of sex. These effects were prevented by combined caffeine intake (0.1 g/L, p.o., ad libitum). Overall, the present study heralds a key role for caffeine intake in the modulation of nandrolone decanoate-induced behavioral changes in rats, suggesting adenosine receptors as candidate targets to manage impact of AAS on brain function and behavior.

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This paper reports the use of paper spray mass spectrometry (PS-MS) combined with chemometric models to analyze seized samples of anabolic steroids. Because many forensic laboratories typically demand high-throughput analysis for this type of sample, we developed a quicker and simpler alternative analytical method for routine analysis with minimal sample preparation. Oily samples (n = 39) resulting from seizures carried out by Brazilian Federal and State Police units were selected for this study. These samples were analyzed by PS-MS in the positive ion mode and full scan (50-1000 m/z), providing spectra containing patterns of the respective active ingredients present in each product. A principal component analysis (PCA) model was built, which discriminated samples mainly according to their active ingredients and allowed to detect and characterize some cases of product counterfeiting. The variable selection method ordered predictors selection was employed jointly with PCA to improve sample cluster separation and to provide model simplification. The final PCA model was built with three principal components and using only 28 spectral variables. This model accounted for 69.82% of the variance and discriminated samples according to their specific active ingredients.

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The illicit use of anabolic androgenic steroids (AAS) as performance-enhancing drugs remains a global issue threatening not only the credibility of competitive sports but also public health due to the well-documented adverse effects they elicit. AAS abuse is not restricted only to professional sports, but also extends to recreational athletes and adolescents as well as in livestock production as growth-promoting agents. Testosterone and nandrolone are among the AAS most frequently exploited. Gas chromatography-mass spectrometry is the reference method for AAS detection, but it is strictly laboratory-based and cannot be performed on-site. The great potential of aptamers in bioanalytical applications and specifically for the development of simple analytical tools suitable for on-site analysis has been extensively documented. In this report, we describe the selection and identification of aptamers binding nandrolone, exhibiting affinity dissociation constants in the low nanomolar range. A label-free colorimetric assay based on gold nanoparticles was developed using one of these novel aptamers for the detection of nandrolone and/or its metabolites. The assay could be deployed for the rapid, on-site, facile and cost-effective screening of samples and provide qualitative visual results with a red to purple/blue color change being indicative of a positive result.

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