Discerning animal-sourced food in diet using isotope analysis of human scalp hair and fingernails.

PURPOSE: Diet-related diseases are advancing as the leading cause of death globally. As self-reporting of diet by patients can be associated with errors, stable isotopes of human tissues can be used to diagnose diseases, understand physiology, and detect change in diet. This study investigates the effect of type and amount of food on the nitrogen and carbon concentration (Nconc and Cconc) and isotopic composition (δ15N and δ13C) in human scalp hair and fingernails. METHODS: A total of 100 residents participated in the study whereas only 74 individuals provided complete diet history. Sixty-six food items majorly available to them were also collected. The Nconc, Cconc, δ15N and δ13C values of human hair, nails and food items were determined. RESULTS: The Nconc, Cconc, δ15N and δ13C values between plant-sourced and animal-sourced food items, as well as human hair and nail tissue were significantly different (p < 0.05). The δ15N value of human tissues was distinct between lacto-vegetarians and omnivores by 0.9‰. The δ15N and δ13C values of human tissues increased by 0.4-0.5‰ with every 5% increase in the consumption of animal protein. CONCLUSIONS: The study helps to demarcate lacto-vegetarians from omnivores, and estimate the percentage of animal protein in diet based on the dual isotope values of human tissues. It also acts as a reference to determine isotopic composition of hair tissue provided the isotope value of nail tissue is known and vice versa.

Occurrence of palladium and platinum in human scalp hair of adolescents living in urban and industrial sites.

Platinum group elements (PGEs) can be naturally found at very low concentrations in the Earth's crust. However, the increasing uses of PGEs in vehicle exhaust catalysts, in addition to some other applications (industry, jewelry, anticancer drugs) cause their anthropogenic emission and dispersion in the environment. The use of human hair samples analysis is considered a suitable biological indicator to assess human occupational and environmental exposure. It is an easily accessible material for individuals or population groups of non-invasive sampling. The aim of this study is a comparative analysis to investigate human hair content of Pd and Pt in adolescents, of both genders, residing near petrochemical plants of Augusta and Gela, in urban area of Palermo, and Lentini as control site (Sicily, Italy). A total of 108 samples were taken from school students (11-14 years old). Hair samples were cleaned, mineralized, and processed for analyses by inductively coupled plasma-mass spectrometry (ICP-MS). The samples from the industrial sites of Gela and Augusta do not have statistically significant differences between them for either Pd for Pt; however, they differ from the samples relating to the city of Palermo. Median Pd concentrations are higher than Pt in industrial sites and control site. In urban site the levels of both metals were comparable. The study does not reveal any statistically significant difference between Pd and Pt concentrations in female and male samples. The data confirm that the study areas are heavily affected by industrial and urban emissions of Pd and Pt, representing a potential hazard to the local population.

Potentially toxic elements in human scalp hair around China's largest polymetallic rare earth ore mining and smelting area.

There is a growing concern about human health of residents living in areas where mining and smelting occur. In order to understand the exposure to the potentially toxic elements (PTEs), we here identify and examine the cadmium (Cd), chromium (Cr), copper (Cu), manganese (Mn), nickel (Ni), lead (Pb) and zinc (Zn) in scalp hair of residents living in the mining area (Bayan Obo, n = 76), smelting area (Baotou, n = 57) and a reference area (Hohhot, n = 61). In total, 194 hair samples were collected from the volunteers (men = 87, women = 107) aged 5-77 years old in the three areas. Comparing median PTEs levels between the young and adults, Ni levels were significantly higher in adults living in the smelting area while Cr was highest in adults from the mining area, no significant difference was found for any of the elements in the reference area. From the linear regression model, no significant relationship between PTEs concentration, log10(PTEs), and age was found. The concentrations of Ni, Cd, and Pb in hair were significantly lower in the reference area when compared to both mining and smelting areas. In addition, Cu was significantly higher in the mining area when compared to the smelting area. Factor analysis (FA) indicated that men and women from the smelting area (Baotou) and mining area (Bayan Obo), respectively, had different underlying communality of log10(PTEs), suggesting different sources of these PTEs. Multiple factor analysis quantilized the importance of gender and location when combined with PTEs levels in human hair. The results of this study indicate that people living in mining and/or smelting areas have significantly higher PTEs (Cu, Ni, Cd, and Pb) hair levels compared to reference areas, which may cause adverse health effects. Remediation should therefore be implemented to improve the health of local residents in the mining and smelting areas.

Elemental Analysis of Laryngeal Cancer Patients in Comparison with Controls Using Scalp Hair as an Analytical Tool.

Laryngeal cancer, a rare form of head and neck cancer, is more common in men, and various factors play important roles in its etiology. In the current study, trace elemental contents in the scalp hair of laryngeal cancer patients and healthy subjects were evaluated. Selected essential and toxic elements were measured using flame atomic absorption spectrophotometry after the hair samples were digested with HNO3-HClO4. The mean/median levels of Pb in the scalp hair of patients were significantly higher than the healthy donors, but Sr, Zn, and Co exhibited comparatively lower levels in the laryngeal cancer patients. In comparison to healthy donors, the correlation pattern among the elements in the hair of the patients exhibited diverse mutual relationships. The multivariate apportionment of the elemental levels in the scalp hair of both donor groups was significantly divergent. The average elemental levels varied significantly among the cancer types (glottic, supraglottic, transglottic, and subglottic) as well as the cancer stages (I, II, III, IV). The average levels of the elements also exhibited considerable variations depending on the donors' habitat, gender, food, and smoking habits. Overall, the study revealed significantly divergent disparities among the elemental contents in the scalp hair of the laryngeal cancer patients in comparison with counterpart controls.

The Attractivity of the Head Louse, Pediculus humanus capitis (Pediculidae: Anoplura) to Isolated Compounds and Blends of Human Skin.

Human head lice Pediculus humanus capitis De Geer (Phthiraptera: Pediculidae) are ectoparasites that cause pediculosis, a global scale disease mainly found in school-age children. Previous works from our laboratory found nonanal, sulcatone, and geranylacetone as the main human scalp volatile components, and individually evaluated their attraction to head lice using an olfactometer. In this work, we compared how their blends at different concentrations attract head lice, and how their blended effect compares to the effect of isolated compounds. At the concentrations evaluated, individual components did not show attraction towards head lice, but a ternary mixture of them was attractive. Moreover, a solvent extract from the human head scalp was analyzed by GC-MS, finding that tetradecanoic acid, palmitic acid, oleic acid, palmitoleic acid, and squalene are the most abundant components. Attraction to these individual compounds at natural concentrations was tested by bioassays in a circular experimental arena. No attraction was observed when the components were tested individually, but when they were evaluated as a blend they attracted head lice. This work presents new information about how chemical signals are attractive at certain concentrations and proportions. This information could be used to better understand communication mechanisms in head lice and for the development of louse repellents.

A quantitative ultra-performance liquid chromatography high-resolution mass spectrometry analysis of steroids from human scalp hair.

Hair steroid analysis offers the possibility to evaluate long-term steroid metabolism. It is especially beneficial when studying the steroid milieu in newborns and children, for example it can provide new insights into steroid metabolism over months and is unaffected by diurnal fluctuations in steroid concentrations. This study describes a quantitative and highly selective high-resolution mass spectrometry method for the simultaneous analysis of multiple steroids from human scalp hair. The developed method utilizes parallel reaction monitoring in the analysis of hydroxylamine derivatized steroids from hair samples first washed with isopropanol, extracted with methanol, and further purified with solid-phase extraction and liquid-liquid extraction. The method was proven suitable for the quantitative analysis of endogenous glucocorticoids (cortisol [1.5-364 pg/mg]; cortisone [3.6-355 pg/mg]; corticosterone [0.7-347 pg/mg]; 11-deoxycortisol [0.1-343 pg/mg]), androgens (testosterone [0.1-288 pg/mg]; dehydroepiandrosterone [0.3-288 pg/mg]; androstenedione [0.1-285 pg/mg]; 11ß-hydroxyandrostenedione [0.3-304 pg/mg]), progestogens (pregnenolone [0.1-313 pg/mg]; progesterone [0.1-313 pg/mg]; 17α-hydroxypregnenolone [0.3-315 pg/mg]; 17α-hydroxyprogesterone [0.7-330 pg/mg]; 21-hydroxyprogesterone [0.6-320 pg/mg]), and estrogens (estrone [0.1-267 pg/mg]; estradiol [0.5-274 pg/mg]). In addition, 11-ketoandrostenedione [0.6-60 pg/mg]; dihydrotestosterone [0.3-290 pg/mg]; 11-ketotestosterone [0.1-12 pg/mg]; and 5α-androstanedione [0.6-281 pg/mg] could be analyzed semi-quantitatively. Aldosterone [3.5-346 pg/mg]; 11ß-hydroxytestosterone [0.3-300 pg/mg]; and 11-ketodihydrotestosterone [0.3-300 pg/mg] were also included in the method, but their concentrations were below the lower limit of quantification in all tested hair samples. The method was applied for the analysis of authentic hair samples from different age groups ranging from newborns to adults, including mothers within 48 h after delivery. The newborn hair samples displayed the widest variety and had also the highest amounts of steroids in comparison to the samples of the other groups.

Direct digestion of human scalp hair on the substrate used for total reflection X-ray fluorescence analysis.

It is important to determine the elemental content of scalp hair to evaluate human health and environmental contamination. Here, a new sample preparation method for total reflection X-ray fluorescence analysis was developed by directly digesting the human hair on the sample holder. A human hair sample was subjected to thermal nitric acid treatment for sample digestion and homogenization. The Zn concentration was estimated to be ~ 1.89 × 102 µg/g. We can evaluate other elements of human hair by using this method.

Interaction Between Essential (Zn) and Toxic (Cd) Elements in Different Stages of Female Breast Cancer Patients, Resident in Different Cities of Sindh, Pakistan.

Breast cancer is the most familiar solid tumor analyzed in women. Trace elements have critical roles in cancer biology. In this research, the relationship between carcinogenic element, cadmium (Cd), and anti-carcinogenic elements, zinc (Zn), in the scalp hair and blood samples of four stages of female breast cancer patients was studied. We have determined the essential trace (Zn) and toxic (Cd) elements, in biological samples (scalp hair and blood) of female breast cancer (n = 96 age ranging 22-35 years), residents of various cities of Pakistan. For comparative study, the biological samples of age-matched healthy (referent) subjects (n = 115) were also analyzed for selected metals. The validity and accuracy of the methodology were checked by using certified reference materials of biological referent materials (human hair (BCR 397) and ClinCheck lyophilized blood). The mean concentrations of Cd were found to be 3- to fourfold significantly higher in the scalp hair and blood samples of female breast patients as compared to referents, while reverse results were obtained in the case of Zn (p > 0.001). The observed data shows the significant effect of carcinogenic (Cd) and their balance towards the anti-carcinogenic (Zn) in humans.

Evaluation of zinc and cadmium levels in the biological samples of Ewing sarcomas patients and healthy subjects.

BACKGROUND: Ewing's sarcoma is a very rare type of cancerous tumor that grows in bones or the soft tissue around the bones, such as cartilage or the nerves. It frequently affects the people at the age of 10 to 20 years and has elevated rate of being cured. METHODS: Assessment of essential trace [zinc (Zn)] and toxic [cadmium (Cd)] elements in biological samples (scalp hair and blood) of Ewing sarcoma patients (n = 87 age ranging 07-19 years) residents of various cities of Pakistan was done. For comparative study, the biological samples of age matched healthy (referent) subjects (n = 62) were also analyzed for selected metals. The matrices of biological samples was oxidized with the help of HNO3 (65%) and H2O2 (30%) mixture at the ratio of 2:1 with the aid of microwave oven. The oxidized biological samples were subjected to atomic absorption spectrometry for their determination. RESULT: The Zn contents in the scalp hair and blood samples of different types of Ewing sarcoma patients were found to be lower, in the range of (45.9-141.2 µg/g) and (0.65-3.12 mg/l), respectively than the biological samples of referent subjects (246-265 µg/g) and (6.40-7.25 mg/l), respectively. Whilst, the Cd concentrations in the scalp hair and blood samples of different types of Ewing sarcoma patients were found to be higher, in the range of (2.70-5.60 µg/g) and (2.46-5.64 µg/l), respectively than the biological samples of controls (1.49-1.79 µg/g) and (1.52-1.90 µg/l), respectively. The biochemical parameters including hemoglobin% and RBC counts were significantly lower in patients than referents (p < 0.001), whereas WBC counts was alarmingly higher in patients than referents (p < 0.001). CONCLUSION: The resulted data will be helpful to treat patients of Ewing sarcoma with mineral supplement contains essential trace element (Zn) in recommended dose to further increase their survival rate.

Assessment of Hg accumulation in fish and scalp hair in fishing communities along river Swat, Pakistan.

Mercury (Hg) bioaccumulation in fish poses severe threats to the food safety and human health. This study was conducted to assess Hg bioaccumulation in fish (n = 24) and scalp hair (n = 77) of the fishing communities at up- and downstream of the river Swat, Pakistan. The mean Hg concentration in upstream fish Salmo trutta fario (Brown trout) and Schizothorax plagiostomus (Swati fish) species was 34.7±18 µg kg-1 and 29.4±15 µg kg-1, respectively. The mean Hg concentration in downstream Swati fish, Crossocheilus diplochilus (Spena deqa), and Garra gotyla (Tora deqa) was 65±21 µg kg-1, 123±33 µg kg-1, and 326±53 µg Kg-1, respectively. The mean Hg concentration in scalp hair of the up- and downstream fishing communities was 658±125 µg kg-1 and 3969±791 µg kg-1, respectively. Independent T-test showed significant difference (p < 0.001) in the mean Hg concentration in scalp hair of the up- and downstream communities. The most prevalent health problems found in the fishing community were muscle pain, headache, visual impairment, arterial blood pressure, anemia, and kidney dysfunction. Multiple linear regression indicated that daily and weekly consumption of the fish significantly increase Hg accumulation in human scalp hair. Regular consumption of fruits and cruciferous and leafy vegetables were found to reduce Hg toxicity in the population. Further studies are recommended to identify the sources of Hg and welfare impact of fish contamination on the fishing community of river Swat.

Adverse impact of occupational exposure on Laborers of cement industry have scalp psoriasis and Pityriasis amiantacea with deficiency of zinc and selenium: impact of mineral supplement.

The relationship between occupational exposure and different skin diseases including scalp psoriasis and Pityriasis amiantacea has been studied. The information about imbalance of essential trace elements in psoriatic patients is still scarce. In the present follow-up study, the scalp hair and blood samples of patients (labors of cement factory) have different skin disorders (plaque's psoriasis and Pityriasis amiantacea) and were analyzed for zinc (Zn) and selenium (Se) before and after treatment with mineral supplements. For comparative purposes, both biological samples of referents who do not have any skin disease (office workers and labors of cement industry) were also analyzed for Zn and Se using an atomic absorption spectrometer. The result indicates a significant decrease in the concentrations of Se and Zn in both biological samples of patients with scalp psoriasis and P. amiantacea as compared with referents/controls, whereas the referent labor has not any skin disease with 15-25% lower values of both essential trace elements as compared to office worker, possibly due to poor diet.

Scalp Hair Metal Analysis Concerning DNA Damage in Welders of Peshawar Khyber Pakhtunkhwa Pakistan.

Welding is used throughout the world in refineries, thermal power plants, chemical facilities, and pressurized containers, and the welders are exposed to toxic heavy metals, electromagnetic fields, polycyclic aromatic hydrocarbon, and ultraviolet radiations. In the present study, 59 welders and an equal number of control subjects were assessed for DNA damage in the lymphocytes using the comet assay. Heavy metals such as lead (Pb), iron (Fe), nickel (Ni), chromium (Cr), manganese (Mn), and cadmium (Cd) levels in the scalp hair of the subjects were evaluated by using atomic absorption spectroscopy (AAS). The results of the current study showed that DNA damage in the lymphocytes of welders (121.8 ± 10.7) was significantly higher as compared with controls (56.5 ± 17.6) (P < 0.001). Besides, the levels of Pb, Fe, Ni, Cr, Mn, and Cd were remarkably higher in the scalp hair of workers as compared with the control group (P < 0.001). Regression analysis showed a prominent association between the heavy metals and total comet score (TCS) in the exposed subjects. Age and duration of occupational exposure had significant effects (P < 0.05) on TCS values. Our results concluded that occupational exposure to welding fumes may cause DNA damage and can lead to important health hazards in the workers.

Primary Cutaneous Myxoid Spindle Cell Squamous Cell Carcinoma of the Scalp: A Case Report.

Cutaneous squamous cell carcinomas are the second most frequent type of non-melanoma skin cancer. A 78-year-old man with a slow-growing but large nodular lesion on his scalp presented to the hospital. The nodular lesion was excised. Histologically, the lesion was diagnosed as a primary cutaneous myxoid spindle cell squamous cell carcinoma, which is the subject of this case report. Extracellular mucin production is an even less common finding in SCC. We also aim to discuss the histological and immunohistochemical findings for distinguishing MSC SCC from other primary cutaneous and metastatic spindle cell neoplasms with myxoid stroma.

Trichilemmal Cysts With Divergent Ductal Differentiation: A Series of 4 Cases.

ABSTRACT: Trichilemmal cysts are common clonal tumors with a predilection for the scalp. They are composed of an outer epithelial wall resembling the outer root sheath in the isthmus of the hair follicle and a central core of compact keratin. Sweat duct differentiation is exceptional with only one convincing case reported to date. Here, we sought to characterize the clinicopathological characteristics of sweat duct differentiation in trichilemmal cysts. We reviewed all cases of trichilemmal cyst diagnosed at our institution between 2008 and 2019. Ductal structures were found in 4 of 411 cases (0.97%). Subjects included 2 male and 2 female patients with a median age of 37.5 years (range 34-55). The ducts were lined by attenuated epithelial cells and immunoreactive for polyclonal carcinoembryonic antigen and cytokeratin 7. Ductal differentiation involved a median of 7.5% (range 1%-50%) of the cyst wall. All 4 cases were from the scalp and treated with local excision. No recurrence was identified with a median follow-up period of 1.5 years (range 1-12 years). In summary, sweat duct differentiation in trichilemmal cysts is rare but likely under recognized. Conceptually, we suggest it represents a type of divergent cellular differentiation within a clonal neoplasm rather than a retention cyst or hybrid cyst.

Fetal scalp blood sampling: Do pH and lactates provide the same information?

OBJECTIVE: Assess the discordance between scalp pH and lactates performed from the same sample during labor. METHOD: This single-center retrospective study included all women with a singleton fetus who had at least one fetal blood sample taken during labor. Some of them had up to seven samples. Scalp pH was the reference parameter for obstetric decision-making. The correlation between the pH and lactates was studied using Pearson coefficient. By categorizing the values as normal, pre-acidosis and acidosis, we were able to estimate agreement with Cohen's kappa coefficient. The frequency of discordance in the categorization and the factors related to it were studied with univariate and multivariable analyses. Cases of severe acidosis at birth (cord pH < 7.00) and cases with acidosis scalp lactates but normal scalp pH were analyzed. RESULTS: We analyzed 480 samples from 268 fetuses among the 2644 deliveries during the study periode. Fetal blood sampling represented 10 % of deliveries. The scalp pH and lactates results were strongly correlated (r=-0.83), but their agreement was only fair (K = 0.36). In 29.4 % of cases, pH and lactates were discordant. Factors related to discordance were meconium-stained fluid, sampling at full dilation and multiple sampling. Six infants (2.2 %) had severe acidosis at birth. Cases' analyses did not allow to conclude severe acidosis could have been avoided using scalp lactates for obstetric decision-making. CONCLUSION: For more than a quarter of the samples, results were discordant between scalp pH and lactates, especially when cervix was full dilated and when the amniotic fluid was meconium-stained. A randomized controlled trial comparing the relevance of each parameter according to the obstetrical situation would be necessary.

Folliculocystic and Collagen Hamartoma: A Distinct Hamartoma Associated With Tuberous Sclerosis Complex.

ABSTRACT: Tuberous sclerosis complex (TSC) is a neurocutaneous disease characterized by cutaneous and extracutaneous hamartomas. Dermatologic evaluation is critical for early diagnosis because mucocutaneous manifestations account for 4 of 11 major and 3 of 6 minor diagnostic criteria. Folliculocystic and collagen hamartoma (FCCH) is a recently described entity associated with TSC. We herein describe the case of a 28-year-old woman with a history of TSC who presented with a scalp lesion present since childhood. Physical examination revealed a solitary, well-circumscribed exophytic tumor over the occipital scalp measuring 9 × 8 cm and covered with comedones and cyst-like structures. Biopsy of the lesion demonstrated thickening of the collagen bundles throughout the dermis, concentric perifollicular and perivascular fibrosis, an increased number of dilated vessels, and keratin-filled cysts lined by the infundibular epithelium. Clinicopathologic correlation was diagnostic for FCCH. The patient was referred for surgical excision. In addition, we review 11 other cases of FCCH previously reported in the literature.

Human volunteer study of the decontamination of chemically contaminated hair and the consequences for systemic exposure.

The decontamination of exposed persons is a priority following the release of toxic chemicals. Efficacious decontamination reduces the risk of harm to those directly affected and prevents the uncontrolled spread of contamination. Human studies examining the effectiveness of emergency decontamination procedures have primarily focused on decontaminating skin, with few examining the decontamination of hair and scalp. We report the outcome of two studies designed to evaluate the efficacy of current United Kingdom (UK) improvised, interim and specialist mass casualty decontamination protocols when conducted in sequence. Decontamination efficacy was evaluated using two chemical simulants, methyl salicylate (MeS) and benzyl salicylate (BeS) applied to and recovered from the hair of volunteers. Twenty-four-hour urinary MeS and BeS were measured as a surrogate for systemic bioavailability. Current UK decontamination methods performed in sequence were partially effective at removing MeS and BeS from hair and underlying scalp. BeS and MeS levels in urine indicated that decontamination had no significant effect on systemic exposure raising important considerations with respect to the speed of decontamination. The decontamination of hair may therefore be challenging for first responders, requiring careful management of exposed persons following decontamination. Further work to extend these studies is required with a broader range of chemical simulants, a larger group of volunteers and at different intervention times.

Load-deformation characteristics of acellular human scalp: assessing tissue grafts from a material testing perspective.

Acellular matrices seem promising scaffold materials for soft tissue regeneration. Biomechanical properties of such scaffolds were shown to be closely linked to tissue regeneration and cellular ingrowth. This given study investigated uniaxial load-deformation properties of 34 human acellular scalp samples and compared these to age-matched native tissues as well as acellular dura mater and acellular temporal muscle fascia. As previously observed for human acellular dura mater and temporal muscle fascia, elastic modulus (p = 0.13) and ultimate tensile strength (p = 0.80) of human scalp samples were unaffected by the cell removal. Acellular scalp samples showed a higher strain at maximum force compared to native counterparts (p = 0.02). The direct comparison of acellular scalp to acellular dura mater and temporal muscle fascia revealed a higher elasticity (p < 0.01) and strain at maximum force (p = 0.02), but similar ultimate tensile strength (p = 0.47). Elastic modulus and ultimate tensile strength of acellular scalp decreased with increasing post-mortem interval. The elongation behavior formed the main biomechanical difference between native and acellular human scalp samples with elastic modulus and ultimate tensile strength being similar when comparing the two.

Cutaneous Adenosquamous Carcinoma of the Scalp With Intestinal Phenotype.

Cutaneous adenosquamous carcinoma is a mixed, squamous and glandular, rare malignant tumor of the skin characterized by a mixed, squamous, and glandular differentiation. Few cases of this tumor have been so far reported, and even fewer have been thoroughly studied by immunohistochemistry. We report here an exceptional case of cutaneous adenosquamous carcinoma which showed immunohistochemically features of intestinal differentiation, namely because of the expression of keratin 20 and CDX2, a marker of gastrointestinal tumors.

Sorption of G-agent simulant vapours on human scalp hair.

Human scalp hair is a biological matrix that can trap chemical vapours from explosives (TNT), drugs (THC) and chemical weapons (yperite). The external contamination of human's hair following exposure to organophosphorus (OP) nerve agent was simulated by model compounds: triethyl phosphate (TEP) and diisopropyl fluorophosphate (DFP). In this work were exposed strands of hair to vapours of TEP and DFP (3 and 7 ppmv) to model sorption kinetics. Sorption isotherms were also investigated at several contamination levels (80-3000 mg min.m-3). OP nerve agent simulants were extracted from hair by soaking in DCM. Raw extracts were analysed in GC-MS/MS to quantify each simulant content in hair. Results were fitted by applying isotherm or kinetic equations. The best model was found to be bimodal first-order, suggesting the co-existence of two different mechanisms of sorption. The best equation to describe OP vapours incorporation on hair was Freundlich model. Thus hair can be used as a passive sensor able to trap chemical G-agents and can also offer valuable information regarding both individual contamination and proof of exposure to chemical weapons.