Introduction

Agates, including moss agate, are commonly found within fractures and cavities of volcanic rocks such as basalt, andesite, and rhyolite1,2,3. The characteristic filamentous texture of moss agate has been examined from both biogenic and inorganic perspectives. Following the work of Liesegang4, interpretations favoring an inorganic origin such as precipitation involving amphibole supergroup species or chlorite group of minerals5,6 became dominant.

However, recent studies by researchers such as Götze et al.1,2, Hofmann et al.7,8, and Watkins et al.9 have reported the presence of filamentous fabrics in opals and chalcedonies from subsurface terrestrial environments. Based on morphological, chemical, isotopic, and literature-based analyses, these studies suggest that some filamentous textures may be of biogenic origin, thereby underscoring their potential significance in the search for past life on Mars.

While these findings challenge the previously accepted hypothesis of an exclusively inorganic origin, additional empirical data have been required to substantiate such claims. In this study, based on the hypothesis that at least some components of green moss agates are of biogenic origin, we have investigated representative specimens from Japan with the aim of demonstrating their biogenic nature.

In Japan, typical occurrences of green moss agate have been documented in such as the Bodai district along the Japan Sea coast of southwestern Japan and the Imakane district in southwestern Hokkaido (Fig. 1). These agates fill cavities within Miocene rhyolitic and andesitic pyroclastic rocks and are believed to have formed through hydrothermal activity subsequent to the eruption of their host volcanic rocks1. The filamentous fabrics observed in the green moss agates from the Bodai and Imakane districts resemble algae or fungi entombed within transparent silica.

Fig. 1
figure 1

Collection sites of moss agate specimens in the Bodai and Imakane districts, along with geological maps of the surrounding area (simplified from Kaseno10; Oka and Katsutoshi11). Moss agate specimens from the Bodai district were collected directly from bedrock outcrops. In contrast, because the bedrock outcrops are covered by dense vegetations, the specimens from the Imakane district were obtained as pebbles, which are considered to have been transported from the areas of pyroclastic rocks.

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In the present study, we conducted a comprehensive analysis of these green moss agates using polarized light microscopy, X-ray powder diffraction (XRD), Raman spectroscopy, elemental distribution mapping via scanning electron microscopy coupled with energy-dispersive X-ray spectroscopy (SEM–EDS), and carbon isotope ratio (δ13C) analysis employing nano-secondary ion mass spectrometry (NanoSIMS). Based on these analytical results, we examine the potential for the filamentous fabrics in green moss agate to be of biogenic origin.

Results and discussions

The filamentous fabrics observed in typical green moss agates exhibit a morphology reminiscent of algae or fungi embedded within transparent silica (Fig. 2).

Fig. 2
figure 2

Stereomicrographs of polished surfaces of representative moss agates from the Bodai and Imakane districts. The filaments, several micrometers in thickness, intertwine to form thicker and more intricate structures. The specimens are deposited in the Toyama Science Museum.

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X-ray powder diffraction data of the samples are tabulated in Table 1 along with reference data of celadonite-1M and glauconite-1M from the ICDD’s Powder Diffraction File (PDF)12. Diffraction peaks typical of these reference data at the d-spacing around 10 Å were not observed due to the influence of the collimator. The Imakane sample showed additional small peak at ~ 4.19 Å, which is ascribed to goethite. Other than that, the main peaks matched well with the reference data. Celadonite is one of clay minerals belonging to celadonite-subgroup of mica group, having ideal chemical composition of KMgFe3+Si4O10(OH)2. Other species of celadonite-subgroup minerals, such as ferroceladonite, also have the same crystal structure with different chemical compositions. There are also polytypes in these minerals, having the same chemical composition with different stacking sequence of the same layer structures. Glauconite is also not a distinct mineral species but name of a series of minerals related to celadonite, characterized by deficiency in interlayer monovalent cation (mainly potassium) and having a high trivalent iron content13. All of these minerals show similar powder XRD patters, meaning our samples are evidently one of “celadonite” in broad sense. Raman spectra of the samples are also consistent with celadonite or glauconite (see “Supplementary Information 2”).

Table 1 Powder X-ray diffraction data of filamentous fabrics in green moss agates from the Bodai and Imakane districts.
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In general, celadonite typically occurs in volcanic rocks14, whereas glauconite is commonly found as granular form within sedimentary rocks15. The filamentous green minerals in moss agates observed in the present study occurred in volcanic rocks and do not have granular forms. Consequently, the minerals exhibiting filamentous fabrics in the Bodai and Imakane districts are hereafter referred to as “celadonite.”

To investigate the origin of the filamentous fabrics while minimizing potential contamination during polishing, moss agate specimens containing filamentous fabrics were fractured. Elemental distributions on the freshly exposed surfaces were subsequently analyzed using analytical scanning electron microscopy (Fig. 3).

Fig. 3
figure 3

Scanning electron microscopy (SEM) images and elemental distribution maps of Si, C, and K on the fractured surfaces surrounding the filamentous fabrics of green moss agate from the Bodai and Imakane districts. White ellipses indicate regions within the filamentous fabrics that lack Si, C, and K, which are interpreted as areas where material was removed during splitting. Red ellipses denote carbonaceous matter (CMs) within the filamentous fabrics. All specimens were cleaned using an ultrasonic washer prior to analysis. The specimens are deposited in the Toyama Science Museum.

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Elemental mapping of the fractured samples obtained using SEM–EDS is presented in Fig. 3. Despite the surface roughness of the fracture planes, the elemental distributions within the filamentous fabric and its surrounding areas are discernible. In both specimens from Bodai and Imakane, the interior regions of the filamentous structures are generally characterized by significantly lower silicon content and elevated concentrations of carbon and potassium relative to the exterior regions. Based on the results of X-ray analyses, these interior regions are inferred to consist of celadonite associated with carbon. Certain areas within the filamentous fabric, indicated by red ellipses, exhibit dense carbon concentrations alongside reduced silicon and potassium contents, suggesting the presence of carbonaceous materials (CMs). Thus, the filamentous fabrics in the green moss agates from the Bodai and Imakane are predominantly composed of celadonite with carbon, with partial inclusions of CMs.

The association of CMs and carbon within the celadonite regions can be interpreted as relics of precursor biogenic matter.

To gain further insights into the formation of celadonite and CMs in the filamentous fabrics, elemental distributions on polished surfaces surrounding the filamentous fabrics of green moss agates from the Bodai and Imakane were mapped using analytical scanning electron microscopy (Fig. 4).

Fig. 4
figure 4

SEM images and elemental distribution maps of Si, C, and K on the polished surfaces around the filamentous fabrics of green moss agate from the Bodai and Imakane districts. Regions within the filamentous fabrics lacking Si, C, and K (white ellipses) are interpreted as areas where material was removed during the polishing process. Red ellipses indicate carbonaceous matter (CMs) within the filamentous fabrics. The specimens were cleaned using an ultrasonic washer. The basic structural relationships between the filamentous fabrics and elemental distributions are consistent with those shown in Fig. 3. Various internal textures (red arrows) within the celadonite of the filamentous fabrics are observable in the SEM images. The specimens are deposited in the Toyama Science Museum.

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Initially, we had concerns regarding potential elemental contamination introduced during surface polishing. However, the fundamental relationship between the carbon distribution and the carbon-rich regions (CMs) within the filamentous fabrics shown in Fig. 4 exhibits no significant difference from that observed on the fractured surface in Fig. 3.

We concluded that substantial contamination during the polishing process was not significant. Consequently, the subsequent discussions are considered valid for the polished surfaces.

To gain a better understanding of the internal textures of the filamentous fabrics, we carefully observed the SEM images of the polished surfaces of the filamentous fabrics (Fig. 5).

Fig. 5
figure 5

SEM images of the insides of filamentous fabrics on the polished planes of green moss agates from Bodai and Imakane Districts. We can recognize the various inner textures on the celadonite in the filamentous fabrics such as the forms of threads (red arrow 1), rings (red arrow 2), cut forms of pipes (red arrow 3) or indeterminate forms (red arrow 4). The specimens are deposited in the Toyama Science Museum.

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The diverse internal textures of the celadonite (Fig. 5) cannot be attributed to contamination during the polishing processes. Similarly, their morphologies should not be interpreted as the crystal habit of celadonite, a monoclinic mineral. Instead, these textures are best understood as relics of biogenic matter precursors predating the precipitation of celadonite.

The CMs within the filamentous fabrics likely represent regions unaffected by corrosion, where celadonite precipitation did not occur. The processes of biogenic matter corrosion and dissolution, coupled with celadonite precipitation, may have taken place when the host volcanic rocks were submerged under seawater, as suggested by Odin14.

To elucidate the mechanism responsible for the formation of the filamentous fabrics, thin sections of these fabrics and adjacent areas in the green moss agates from the Bodai and Imakane districts were examined using polarized light microscopy (Fig. 6).

Fig. 6
figure 6

Polarized light micrographs of filamentous fabrics in green-colored moss agates from the Bodai and Imakane districts, observed under plane-polarized light and crossed polars. Dark areas visible under plane-polarized light (indicated by red ellipses) correspond to carbonaceous matter (CMs). The specimens are housed in the Toyama Science Museum.

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In Fig. 6, filamentous fabrics exhibiting green to yellow-green hues (mainly celadonite) are enclosed by amorphous opal grains with a low refractive index and displaying a pale pink coloration. Both the filamentous fabrics and the opal are subsequently embedded within radially extinct fibrous chalcedony. The chalcedony is identified as length-fast common chalcedony (see “Supplemental Information 3”).

The opal grains exhibit rounded morphologies against the outer chalcedony boundary and straight interfaces against the inner filamentous fabrics. Opal grains with rounded forms at the time of precipitation tend to adhere to pre-existing fabrics16. The morphology of the opals observed in Fig. 6 provides compelling evidence that the opal grains adhered to the filamentous fabrics prior to the formation of chalcedony. Subsequently, chalcedony precipitated, filling the remaining voids.

Based on these observations, the formation sequence of the observed fabrics is interpreted as follows:

  1. 1.

    Formation of filamentous fabrics in the hot spring environment,

  2. 2.

    Precipitation of opal onto the filamentous fabrics, and

  3. 3.

    Subsequent precipitation of chalcedony, encapsulating both the pre-existing filamentous fabrics and opal.

These conclusions are consistent with the findings of Akahane et al.17.

The origin of the carbon determined from the isotope ratio has been broadly discusse18,19,20,21,22,23,24. As a result, carbon having an isotope ratio (δ13C) higher than − 16‰ would be considered as representing an “inorganic matter origin”, while an isotope ratio of less than − 12‰ would be considered as representing an “biogenic matter origin” (Fig. 7).

Fig. 7
figure 7

Carbon isotope ratios (δ13C) of carbonaceous materials (CMs) within the filamentous fabrics of green moss agates from the Bodai and Imakane districts, analyzed using NanoSIMS. Numerical values (left) and corresponding graphical representations are presented. The limits of the labels “biogenic origin” and “inorganic origin” are based on previously reported literatures18,19,20,21,22,23,24. Imakane1_1 and Imakane1_2, and Imakane2_1 and Imakane2_2 were repeated analyses of the same specimens respectively. All specimens are deposited in the Toyama Science Museum.

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We have 15 analyses of carbon isotope ratios (δ13C) of CMs in filamentous fabrics of green moss agates from the Bodai and Imakane in total. Eleven analyses (BodaiA_2, BodaiA_3, BodaiA_4, BodaiC_3, BodaiC_5, BodaiC_6, ImakaneB_1, Imakane1_1 = Imakane1_2 and Imakane2_1 = Imakane2_2) of 9 specimens were in the area denoted as being of “biogenic origin” (Fig. 7). The CMs in those filamentous fabrics can be considered as biogenic matter origin.

On the other hand, the Bodai 1 and Bodai 2 specimens (Fig. 7) exhibit large errors that span both the inorganic and biogenic regions. Although the Bodai A_1 and Bodai C_4 specimens display smaller errors, their values still straddle the boundary between the biogenic and inorganic regions (Fig. 7). Consequently, it is not possible to definitively determine whether the isotope ratios of Bodai 1, Bodai 2, Bodai A_1, and Bodai C_4 are of biogenic or inorganic origin. Nevertheless, of the 15 measured carbon isotope ratios, the remaining 11 values fall within the biogenic region.

Based on this evidence, we conclude that, although some contamination may have occurred during polishing or other preparation processes, the majority of the CMs from the Bodai and Imakane are likely of biogenic origin. In other words, most of the CMs from these districts are considered to be relics of the precursor biogenic matter from which the celadonite originated.

Conclusion

The typical green moss agates found in the volcanic rocks of the Bodai and Imakane exhibit filamentous fabrics that are only a few microns thick and become intertwined, forming thicker aggregates (Fig. 2). Opal particles adhere to the surfaces of these filaments, and both the filaments and opal particles are embedded within transparent chalcedony (Fig. 6). Powder XRD and Raman spectroscopy indicate that the filaments primarily consist of celadonite or glauconite, with minor amounts of goethite. Since distinguishing between celadonite and glauconite is highly challenging, a definitive identification was not possible through analysis alone. However, considering its occurrence in volcanic rocks and morphology, we identified it as celadonite. SEM–EDS mapping (Figs. 3, 4) on both fractured and polished surfaces revealed that the interior of the celadonite filaments is enriched in carbon compared to the outer regions, with certain areas exhibiting a high concentration of CM. No significant elemental distribution differences were observed between the fractured (Fig. 3) and polished (Fig. 4) surfaces under EDS mapping. Various internal textures of the celadonite filaments (Fig. 5) suggest that they are not contaminants introduced during the polishing process or mineralogical growth features of celadonite. Instead, these structures are likely remnants of biogenic precursor materials that had existed before celadonite precipitation. Carbon isotope ratio analyses of the CM within the filaments indicated biogenic origins for the majority (11 out of 15 spots), while a few spots (4 out of 15) might be affected by surface contamination. These results suggest that the celadonite filaments originally formed through the corrosion and replacement of biogenic precursors, with the carbonaceous matters (CM) representing remnants that survived these processes. The most probable biogenic precursors are biological organisms such as fungi or algae from the viewpoint of their size. The probable formation sequence of filamentous fabrics in green moss agate follows the stages: (1) formation of filamentous fabrics in hydrothermal water, (2) precipitation of opal into the filaments, and (3) deposition of chalcedony, embedding the pre-existing filaments and opal.

We conclude that most filamentous fabrics observed in the green moss agate examined in this study are of biogenic origin. This finding, supported by both morphological and geochemical evidence, provides compelling confirmation of previous claims that at least some filamentous fabrics observed in subsurface environments are of biogenic origin, as proposed by Götze et al.1,2, Hofmann et al.7,8, and Watkins et al.9.

Future research is expected to confirm the biogenic origin of filamentous fabrics in green moss agate from various geological periods, further enhancing our understanding of Earth’s history through their paleo biological significance.

Methods

As shown in Fig. 1, Specimens from the Bodai District were collected from the bedrock composed of Miocene rhyolite lavas and pyroclastic rocks. In contrast, specimens from the Imakane District were gathered as pebbles derived from Miocene andesitic pyroclastic rocks (Pyroclastic Member of the Kunnui Formation). Two of the polished samples used in Table 1 are deposited in the collection of the National Museum of Nature and Science, Tokyo. The registration number of the sample from Bodai is NSM-M53369, and that of Imakane is NSM-M53368. All other specimens used in Figs. 2, 3, 4, 5, 6 and 7 are deposited in the Toyama Science Museum (see “Supplementary Information 1”).

The specimens were polished, and stereomicrographs were taken under floodlighting from an obliquely upward direction (Fig. 2). Thin sections of the specimens (Fig. 6) were polished on both sides, mounted on slide glass using epoxy resin, and examined under polarized light microscopy.To obtain elemental distribution maps of the specimens, analytical scanning electron microscopy (JEOL, JSM-6510LA) was performed at the Fossa Magna Museum, as shown in Figs. 3, 4, and 5. The analyses were conducted under low-vacuum conditions (50–80 Pa) with an accelerating voltage of 15 kV. Elemental analysis and mapping were carried out using an energy-dispersive X-ray spectroscopy (EDS) system (JEOL, EX-54185JMU) integrated with the scanning electron microscope. The qualitative intensities of each element are indicated by the scale bar located at the lower left of each image.

NanoSIMS Ion Imaging Analysis: The carbon isotopic compositions of the samples were determined using isotopic imaging with a NanoSIMS 50L instrument (Ametek CAMECA) at JAMSTEC. This analysis provided isotope map of carbon (13C/12C ratio or d13C) and elemental maps of 16O, 12C2, and 12C14N. In briefly, a focused primary Cs+ beam with a current of approximately 2 pA was rastered over regions of interest (48 × 48 μm2). Each measurement was preceded by presputtering with a high primary ion beam current to stabilize the secondary ion beam intensity. For carbon and nitrogen isotopic analysis, images of 12C−, 13C−, 16O−, 12C2−, 12C14N− were acquired simultaneously by multidetection with five electron multipliers at a mass resolving power of approximately 9000, which was sufficient to separate all relevant isobaric interferences (that is, 12C1H on 13C). Subsequent analyses involved ten repeated scans over the same area, producing images with a resolution of 256 × 256 pixels. The dwell time was 1000 μs per pixel, resulting in a total acquisition time of approximately two hours per analysis. Instrumental mass fractionation was corrected using the USGS24 graphite standard as a reference for carbon isotope measurements. Although the carbon isotope fractionation factors varied on a daily basis, the reproducibility within each analytical session was within 1–2‰. Detailed analytical procedures of carbon isotopic compositions for materials were published elsewhere25,26,27.

Powder X-ray diffraction (PXRD) data were acquired using a Gandolfi camera with a diameter of 114.6 mm and Ni-filtered CuKα radiation (λ = 1.54187 Å) at the National Museum of Nature and Science. The diffraction patterns were recorded on an imaging plate and processed with a Fuji BAS-2500 bio-image analyzer and software developed by Nakamuta14.

Raman microscopy was performed using a Raman spectrophotometer (JASCO, NRS-7100) that was equipped with a 532 nm semiconductor green laser the Division of Instrumental Analysis, University of Toyama. Laser power was obtained at ca. 13 mW on the sample surface. The peak position of silicon wafer (520.7 cm−1) was used for calibration of the Raman band position. Raman spectra of each analytical spots were obtained by accumulating 10 acquisitions of 3 s, using a 1800 grooves/mm grating and a CCD camera (1024 × 256 pixels), where pixel-spectral resolution becomes approximately 0.8 cm−1.