Prevendar Crnić et al. 2022 — Hazardous trace elements in bird tissues, Raša coal area, Croatia
This study measured 13 elements (Al, V, Cr, Mn, Fe, Co, Cu, Zn, As, Se, Mo, Cd, Pb) by ICP-MS in muscle, liver, kidney, and brain of 12 wild birds (pigeon, jay, mallard, magpie) from the Raša coal-polluted area of eastern Istria, Croatia and 8 control-area birds (pigeon, pheasant, jay) from the Kontija Forest ~35 km west of the pollution source. Of the 13 elements, the authors report distributions for 5 (Se, Cd, Pb, Cu, Zn) selected for elevated occurrence in the polluted environment. Mean Se and Cd were significantly elevated in Raša-area birds vs control (Se muscle p=0.003, Se liver p=0.04, Cd kidney p=0.01 by Mann–Whitney U). Highest Cu, Zn, and Se concentrations occurred in liver; highest Cd and Pb in kidney. Mallard kidney Cd (mean 2.30 mg/kg w.w.) and pigeon kidney Cd (1.43 mg/kg w.w.) exceed the Croatian Ordinance 146/12 maximum allowable Cd in poultry kidney (1.0 mg/kg w.w.) used as a reference standard because no game-bird-specific Croatian Cd threshold exists. The authors nonetheless conclude human-health risk from consumption of these game species is very low because mean annual game-meat consumption in Croatia is only 0.3 kg per household member (Croatian Statistical Yearbook 2011).
Key numbers
Table 1 — Pooled-species Raša (R, n=12) vs Control (C, n=8) descriptive statistics by tissue (mg/kg w.w.; brain C n=6 for all metals, brain R n=11 and brain C n=6 for Pb specifically, kidney C n=7 for Pb specifically):
| Metal · Tissue · Area | n | Mean | Median | Min | Max | Q1 | Q3 | SD |
|---|---|---|---|---|---|---|---|---|
| Cu muscle R | 12 | 5.27 | 5.08 | 4.09 | 6.98 | 4.48 | 5.86 | 0.972 |
| Cu muscle C | 8 | 3.91 | 4.01 | 0.762 | 6.34 | 3.27 | 4.80 | 1.64 |
| Cu liver R | 12 | 17.7 | 4.37 | 2.74 | 68.1 | 3.32 | 234 | 25.7 |
| Cu liver C | 8 | 3.76 | 3.29 | 2.86 | 6.13 | 3.17 | 4.10 | 1.06 |
| Cu kidney R | 12 | 4.76 | 4.21 | 3.56 | 7.28 | 3.95 | 5.41 | 1.31 |
| Cu kidney C | 8 | 4.13 | 4.09 | 3.26 | 5.30 | 3.59 | 4.54 | 0.685 |
| Cu brain R | 12 | 2.89 | 2.77 | 2.33 | 4.58 | 2.49 | 3.02 | 0.592 |
| Cu brain C | 6 | 3.15 | 3.10 | 2.37 | 4.26 | 2.80 | 3.25 | 0.627 |
| Zn muscle R | 12 | 10.6 | 10.3 | 7.80 | 14.3 | 9.76 | 11.7 | 1.81 |
| Zn muscle C | 8 | 9.71 | 9.29 | 6.65 | 14.9 | 8.71 | 10.1 | 2.36 |
| Zn liver R | 12 | 34.0 | 30.0 | 20.3 | 68.4 | 24.5 | 41.4 | 14.1 |
| Zn liver C | 8 | 26.8 | 25.6 | 19.2 | 41.0 | 19.6 | 32.0 | 7.91 |
| Zn kidney R | 12 | 25.7 | 25.7 | 21.6 | 31.3 | 23.1 | 28.1 | 2.99 |
| Zn kidney C | 8 | 26.8 | 27.7 | 20.7 | 32.3 | 22.9 | 30.1 | 4.32 |
| Zn brain R | 12 | 12.1 | 12.2 | 10.0 | 14.5 | 10.8 | 13.2 | 1.40 |
| Zn brain C | 6 | 12.0 | 11.6 | 10.1 | 14.6 | 11.2 | 12.9 | 1.54 |
| Se muscle R | 12 | 0.776 | 0.406 | 0.150 | 3.36 | 0.308 | 0.939 | 0.926 |
| Se muscle C | 8 | 0.196 | 0.192 | 0.114 | 0.301 | 0.150 | 0.236 | 0.0623 |
| Se liver R | 12 | 1.78 | 0.721 | 0.230 | 7.51 | 0.544 | 1.87 | 2.34 |
| Se liver C | 8 | 0.517 | 0.450 | 0.198 | 1.16 | 0.322 | 0.616 | 0.303 |
| Se kidney R | 12 | 1.67 | 1.14 | 0.598 | 4.60 | 0.973 | 1.84 | 1.24 |
| Se kidney C | 8 | 0.947 | 0.952 | 0.552 | 1.36 | 0.696 | 1.18 | 0.292 |
| Se brain R | 12 | 0.398 | 0.272 | 0.177 | 0.971 | 0.237 | 0.513 | 0.274 |
| Se brain C | 6 | 0.223 | 0.235 | 0.180 | 0.258 | 0.181 | 0.249 | 0.0346 |
| Cd muscle R | 12 | 0.00592 | 0.00508 | 0.000784 | 0.0174 | 0.00303 | 0.00644 | 0.00482 |
| Cd muscle C | 8 | 0.0127 | 0.00290 | 0.000825 | 0.0815 | 0.00202 | 0.00470 | 0.0278 |
| Cd liver R | 12 | 0.389 | 0.319 | 0.0816 | 1.578 | 0.153 | 0.441 | 0.402 |
| Cd liver C | 8 | 0.177 | 0.160 | 0.0525 | 0.417 | 0.124 | 0.190 | 0.108 |
| Cd kidney R | 12 | 1.60 | 1.50 | 0.286 | 4.99 | 0.723 | 1.73 | 1.28 |
| Cd kidney C | 8 | 0.477 | 0.463 | 0.0801 | 0.940 | 0.344 | 0.590 | 0.253 |
| Cd brain R | 12 | 0.00262 | 0.00217 | 0.000479 | 0.00703 | 0.000926 | 0.00346 | 0.00220 |
| Cd brain C | 6 | 0.00167 | 0.00155 | 0.000481 | 0.00315 | 0.000701 | 0.00255 | 0.00106 |
| Pb muscle R | 12 | 0.0364 | 0.00358 | 0.00131 | 0.398 | 0.00204 | 0.00535 | 0.114 |
| Pb muscle C | 8 | 0.0396 | 0.00292 | 0.00158 | 0.292 | 0.00199 | 0.00679 | 0.102 |
| Pb liver R | 12 | 0.232 | 0.0272 | 0.00886 | 2.44 | 0.0217 | 0.0407 | 0.696 |
| Pb liver C | 8 | 0.0666 | 0.0142 | 0.00408 | 0.430 | 0.0116 | 0.0235 | 0.147 |
| Pb kidney R | 12 | 0.249 | 0.109 | 0.0275 | 1.14 | 0.0512 | 0.306 | 0.337 |
| Pb kidney C | 7 | 0.250 | 0.0495 | 0.0212 | 1.45 | 0.0233 | 0.0797 | 0.532 |
| Pb brain R | 11 | 0.0138 | 0.0106 | 0.00117 | 0.0424 | 0.00481 | 0.0200 | 0.0116 |
| Pb brain C | 6 | 0.0156 | 0.00716 | 0.00490 | 0.0606 | 0.00524 | 0.00851 | 0.0221 |
(Internal-mirroring caveat in source Table 1: the “Cu liver R” row gives mean 17.7, Q1 3.32, Q3 234 — the Q3 of 234 mg/kg w.w. is clearly mislabelled in the source given that the column max is 68.1 and the Q1 is 3.32; the published value is reproduced verbatim. See Verification notes.)
Table 2 — Per-species mean concentrations in Raša-area pigeon, mallard, and magpie (mg/kg w.w.):
| Species · Tissue | Cu | Zn | Se | Cd | Pb |
|---|---|---|---|---|---|
| Pigeon · muscle | 5.12 | 10.03 | 0.344 | 0.00450 | 0.00350 |
| Pigeon · liver | 3.56 | 25.7 | 0.575 | 0.231 | 0.0238 |
| Pigeon · kidney | 3.87 | 26.8 | 1.01 | 1.43 | 0.0728 |
| Pigeon · brain | 2.78 | 12.9 | 0.233 | 0.00270 | 0.0158 |
| Mallard · muscle | 6.56 | 12.4 | 2.09 | 0.00360 | 0.00360 |
| Mallard · liver | 59.0 | 53.7 | 5.29 | 0.757 | 0.0410 |
| Mallard · kidney | 6.85 | 23.7 | 3.55 | 2.30 | 0.201 |
| Mallard · brain | 3.56 | 10.6 | 0.840 | 0.00270 | 0.0151 |
| Magpie · muscle | 4.39 | 9.65 | 0.324 | 0.00290 | 0.00360 |
| Magpie · liver | 4.88 | 33.3 | 0.747 | 0.132 | 0.0294 |
| Magpie · kidney | 4.43 | 21.9 | 1.30 | 0.310 | 0.632 |
| Magpie · brain | 2.33 | 12.4 | 0.299 | 0.00100 | 0.0106 |
(Jay per-species means are reported in Table 3 of the source, not Table 2.)
Table 4 — Mann–Whitney U test p-values, Raša vs control area, all-bird pooled (bold = significant at p < 0.05):
| Tissue | Cu | Zn | Se | Cd | Pb |
|---|---|---|---|---|---|
| Muscle | 0.05 | 0.13 | 0.003 | 0.37 | 0.79 |
| Liver | 0.26 | 0.15 | 0.04 | 0.13 | 0.11 |
| Kidney | 0.37 | 0.51 | 0.15 | 0.01 | 0.19 |
| Brain | 0.2 | 0.96 | 0.07 | 0.6 | 0.7 |
Table 5 — Croatian Ordinance 146/12 maximum permitted concentrations applied as reference standards (no game-bird-specific values exist; poultry values used by the authors as the closest applicable reference):
| Tissue (mg/kg w.w.) | Pb | Cd |
|---|---|---|
| Muscle | 0.1 | 0.05 |
| Liver | 0.5 | 0.5 |
| Kidney | 0.5 | 1.0 |
Authors’ fitness-for-consumption comparison (page 27–28):
- Mallard liver Cd mean (0.756 mg/kg w.w.) exceeds the Ordinance 146/12 poultry-liver Cd limit of 0.5 mg/kg w.w.
- Pigeon kidney Cd mean (1.43 mg/kg w.w.) and mallard kidney Cd mean (2.30 mg/kg w.w.) exceed the Ordinance 146/12 poultry-kidney Cd limit of 1.0 mg/kg w.w.
- All-bird-pooled mean Pb in Raša-area muscle (0.0364), liver (0.232), kidney (0.249), and brain (0.0138 mg/kg w.w.) are below the Croatian reference thresholds and below the Franson & Pain (2011) 2 mg/kg w.w. liver/kidney bird-poisoning threshold and the Kalisinka (2000) 1.5 mg/kg w.w. brain Pb poisoning threshold.
- Authors’ conclusion: human-health risk from consumption of meat and offal of these species in the study area is “very low,” because average annual game-meat consumption in Croatia is 0.3 kg per household member (Croatian Statistical Yearbook 2011) and large-game species (deer, wild boar, roe deer) are consumed more than feathered game (ducks, pigeons).
Methods (brief)
Sampling: 20 wild birds collected December 2019 – January 2020 in eastern Istria, Croatia. Raša coal-polluted area (n=12): 5 wild pigeons and 2 mallards from the mouth of the Raša River, 1 pigeon from Koromačno, 1 mallard from Polje Čepić, 1 magpie from Brgod, and 2 jays from Gora Glušići. Control area (n=8): Kontija Forest above the Lim Channel on the west coast of Istria (~35 km west of the pollution source), 6 wild pigeons plus pheasants and jays.
Sample prep: 0.5 g homogenised tissue was weighed into a Teflon liner with 3 mL H₂O + 2.5 mL HNO₃ (65%). High-pressure microwave digestion (Multiwave 3000, Anton Paar, Graz, Austria) in three steps: (I) 5 min at 500 W, (II) 20 min at 1000 W, (III) 30 min at 1200 W. Cooled, transferred to a 50 mL volumetric flask, filled to mark with ultra-pure water. Internal-standard mixture (In, Bi, Sc; Inorganic Ventures, Blacksburg, VA, USA) added on-line via the standard ISTD mixing tee.
Instrumentation: Agilent ICP-MS Model 7900 (Agilent, Palo Alto, CA, USA), high-purity Ar (99.999%, White Martins, Brazil). Calibration using certified Environmental Calibration Standards at 99.999% purity for Se, Cd, Pb, Cu, Zn at 10 mg/L (Agilent Technologies, USA).
Statistical analysis: Descriptive statistics (mean, median, min, max, Q1, Q3, SD) by tissue × area. Mann–Whitney U test for Raša vs control area comparisons; significance level p < 0.05.
Speciation: total arsenic and total mercury are not separately reported in the paper’s results; the 5 elements selected for detailed reporting (Se, Cd, Pb, Cu, Zn) are reported as total. The metals field uses Cd, Pb, Cu, Zn, Se without speciation tags because these elements either do not require speciation (Cu, Zn) or were measured as total concentration (Cd, Pb, Se as total Se).
Basis: all values expressed in mg/kg wet weight (w.w.). Where literature comparisons in Table 2 of the source are drawn from studies reporting dry weight, the authors recalculated those values to w.w. assuming 70% water content in bird tissue (Table 2 caption).
Implications
Certification: Game-meat threshold setting is not currently a high-priority HMTc row, and no game-bird-specific maximum levels exist under Croatian or EU food law (the Croatian Ordinance 146/12 covers cattle, sheep, pigs, poultry, and horses only; the EU Regulation 1881/2006 / 2023/915 successor covers meat and edible offal of livestock without a game-bird subcategory). This paper documents that under coal-pollution context, mallard kidney and pigeon kidney Cd routinely exceed the poultry-kidney reference threshold of 1.0 mg/kg w.w. (mean 2.30 and 1.43 mg/kg w.w. respectively), and mallard liver Cd exceeds the poultry-liver reference threshold of 0.5 mg/kg w.w. (mean 0.756 mg/kg w.w.). For routing into the game-meats HMTc row, the kidney-offal Cd signal is the dominant risk pathway in this dataset.
Courses: Useful as a case study in (1) bioindicator design — birds as longest-best-studied bioindicators of environmental pollution; tissue choice (kidney/liver for long-term exposure, blood for short-term, bone for cumulative); (2) the distinction between essential trace elements (Cu, Zn, Se) showing adequate-to-high but not poisoning-range concentrations in liver and the non-essential trace elements (Cd, Pb) accumulating in kidney; (3) source attribution — Se and Cd are the elements the surrounding superhigh-organic-sulphur Raša coal characteristically enriches (Medunić et al. 2016, 2018a, 2020, 2021), and Se and Cd are the elements with significant Raša-vs-control differences in this study; (4) the species-mix effect — mallards (waterfowl, aquatic-food diet) show systematically higher Se and Cd than pigeons/jays/magpies (terrestrial diet) because Se and Cd bioaccumulate rapidly in aquatic sediments and food webs.
App: Provides Croatian wild-bird occurrence data for the game-meats product page and the meat-and-poultry / organ-meats ingredient pages. Per-species, per-tissue concentration means are reported for Cu, Zn, Se, Cd, Pb for pigeon, mallard, and magpie (Table 2 above) and Raša-vs-control descriptive statistics for the pooled-species sample (Table 1 above). Sampling year range 2019–2020 anchors this dataset toward the recent end of the wild-bird literature.
Microbiome: Not addressed.
Verification notes
- 2026-05-29 (Claude Opus 4.7, ingest-next-manual-fetch-pdf v2.0 — Phase 1): NEW path source page written from the auto-fetched PDF in
raw/manual-fetch/seasonal-geographic-variance/auto-fetched/. Three identity checks (DOI grep on 10.17794/rgn.2022.1.3, raw_handle grep on the auto-fetched filename, cite-key grep for prevendar/crnic/2022/rasa patterns) all negative; no existing wiki page. - Cite-key: first author’s compound surname is “Prevendar Crnić” (Croatian convention). The cite-key uses the hyphenated form
prevendar-crnic2022-...for consistency with other compound-surname pages in the corpus. - Speciation: paper measures total Se, total Cd, total Pb, total Cu, total Zn — no speciation reported for Se (no organic-Se vs inorganic-Se), Cd, or Pb; Cu and Zn do not require speciation in food-occurrence work. The metals field uses bare element symbols accordingly. The paper analytically measured 8 additional elements (Al, V, Cr, Mn, Fe, Co, As, Mo) but did not report distributions; those elements are not added to the metals field because the paper reports no extractable numerical values for them. Notably the paper measured tAs by ICP-MS but did not report any As distribution and did not speciate, so neither iAs nor tAs is added to the metals field.
- Internal-mirroring caveat on source Table 1: the “Cu liver R” row column “Upper quartile (Q3)” reads 234 mg/kg w.w., which is implausible given the column maximum of 68.1 mg/kg w.w., the column Q1 of 3.32 mg/kg w.w., and the column median of 4.37 mg/kg w.w. This appears to be a typesetting error in the source (likely intended 23.4 or 2.34). The published value is reproduced verbatim in the Key numbers table above; no correction is applied because that would be inferring rather than reporting. The same row’s mean of 17.7 mg/kg w.w. is high but plausible because the Raša pooled-species sample includes mallards whose per-species mean Cu liver concentration is 59.0 mg/kg w.w. (Table 2), an outlier driver of the pooled mean.
- Second internal-mirroring caveat: the source reports mallard liver Cd as 0.757 mg/kg w.w. in Table 2 (page 23) and as 0.756 mg/kg w.w. in the narrative text on page 28 (“mallard’s liver (0.756 mg/kg, w.w.)”). Both values are preserved verbatim where the source uses each — 0.757 in the Table 2 reproduction above and 0.756 in the “Authors’ fitness-for-consumption comparison” bullet — and the audit subagent (2026-05-29, ⚠️ advisory) flagged the mismatch as a source-internal inconsistency, not a wiki-side error. No correction applied; the difference is at the third significant figure and does not change the regulatory comparison (both exceed the Ordinance 146/12 poultry-liver Cd limit of 0.5 mg/kg w.w.).
- 2026-05-29 (Claude Opus 4.7, ingest-next-manual-fetch-pdf v2.0 — Phase 3 audit application): fresh-context audit subagent (Phase 2) returned PROMOTE with one ⚠️ advisory presentation finding (the mallard-liver-Cd 0.756/0.757 mismatch noted above) and four ✅ clean checks (slug vocabulary, speciation/methods, Part 12 brand firewall, Part 2 wiki/HMTc firewall). Full Table 1 (40 rows × 8 columns), Table 2 (12 rows × 5 metals), Table 4 (20 cells), Table 5 (6 cells) reproductions all returned verbatim-clean against the PDF. Applied: one verification note added documenting the source-internal 0.756 vs 0.757 mismatch (no numerical values changed in the page). Rejected: none.
- Tables 1 and 2 are reproduced as published; species-mix asymmetry between Raša and control (12 birds Raša pigeon+mallard+jay+magpie vs 8 birds control pigeon+pheasant+jay) is preserved without re-pooling.
- Brand firewall (Part 12): the source names no commercial brand of game meat or supplement. Instrument vendors named in the methods section (Anton Paar, Agilent Technologies, Inorganic Ventures, White Martins) are preserved per the 2026-05-17 scientific-method vendor-name exception.
- Frontmatter slug check: ingredients
meat-and-poultry✓,organ-meats✓; productsgame-meats✓ (HMTc Category 11 Row 5); metals abbreviations Cd, Pb, Cu, Zn, Se all valid (per CLAUDE.md Part 14 metal-vocabulary section, plain element symbols are correct for Cu, Zn, Se; Cd and Pb are also standard); jurisdictions HR ✓ (used elsewhere in the corpus), EU ✓ (EFSA/EU regulatory context is invoked implicitly via Croatian implementation of EU contaminant law). Matricesgame-meat✓ (used by siblinggasparik2017-wild-boar-slovakia-metals), bird-muscle/bird-liver/bird-kidney/bird-brain consistent with corpus bird-tissue convention (verified viagrep -rh '^matrices:' wiki/sources/). - Croatian regulations cited in the source body — Ordinance 146/12 (Pravilnik o najvećim dopuštenim količinama određenih kontaminanata u hrani) and Hunting Act OG 99/18, 32/19, 32/20 (Zakon o lovstvu) — are referenced in Implications as the authors’ applied reference thresholds. They are not added as separate
[[regulations/...]]wikilinks because no Croatian regulation pages exist yet in the wiki; per CLAUDE.md Part 10, regulation pages should be created on first encounter with the correct rule ID, which is a separate workflow from per-paper ingest. This source is flagged as a potential trigger for that workflow but no page is created here. - Wet-weight basis is explicit throughout the paper (all data expressed in mg/kg w.w.). Literature comparisons in source Table 2 were recalculated from dry weight to w.w. by the source authors assuming 70% water content in bird tissue; downstream comparators should preserve that conversion provenance.
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Page history
The five most recent substantive edits to this page. The full version history lives in git; when DOI minting comes online (see schema docs), each entry below will also link to a version-pinned DataCite DOI.
| Commit | Date | Description |
|---|---|---|
| 12d44c4 | 2026-05-29 | audit: eaglessmith2016-freshwater-fish-mercury-western-us-canada [revised] |