Long-term Impact of Gold and Platinum on Microbial Diversity in Australian Soils.

The effects of platinum (Pt) and gold (Au) and on the soil bacterial community was evaluated in four different Australian soil types (acidic Burn Grounds (BGR), organic matter-rich Fox Lane, high silt/metal Pinpinio (PPN), and alkali Minnipa (MNP) spiked with either Pt or Au at 1, 25, and 100 mg kg-1 using a next-generation sequencing approach (amplicon-based, MiSeq). Soil type and metal concentrations were observed to be key drivers of Pt and Au effects on soil microbial community structure. Different trends were therefore observed in the response of the bacterial community to Pt and Au amendments; however in each soil type, Pt and Au amendment caused a detectable shift in community structure that in most samples was positively correlated with increasing metal concentrations. New dominant groups were only observed in BGR and PPN soils at 100 mg kg-1 (Kazan-3B-28 and Verrucomicrobia groups (BGR, Pt) and Firmicutes and Caldithrix groups (PPN, Pt) and WS2 (BGR, Au). The effects of Pt on soil microbial diversity were largely adverse at 100 mg kg-1 and were pronounced in acidic, basic, and metal/silt-rich soils. However, this effect was concentration-related; Au appeared to be more toxic to soil bacterial communities than Pt at 25 mg kg-1 but Pt was more toxic at 100 mg kg-1. More bacterial groups such as those belonging to Burkholderiales/Burkholderiaceae, Alicyclobacillaceae, Rubrobacteraceae, Cytophagaceae, Oxalobacteraceae were selectively enriched by Pt compared to Au (Sphingomonadaceae and Rhodospirillaceae) amendments irrespective of soil type. The research outcomes have important implications in the management (remediation) of Pt- and Au-contaminated environments.

The application of Marinobacter hydrocarbonoclasticus as a bioaugmentation agent for the enhanced treatment of non-sterile fish wastewater.

Wastewaters generated by fish processing are characterised by salt concentrations similar to or greater than that of seawater together with high nutrient concentrations (e.g. organic carbon and total nitrogen) due to the presence of blood, oil, and fish tissues. Fish processing wastewater entering rivers and oceans have become a key factor leading to the pollution of receiving waters; the adequate treatment of this wastewater is, therefore, crucial to a sustainable fish industry. The present study aimed to determine whether augmentation of fish wastewater with either Marinirhabdus sp., Marinobacter hydrocarbonoclasticus or a consortium of the two halobacteria, could successfully enhance the removal of both chemical oxygen demand (COD) and total nitrogen (TN) from fish wastewater. Following 9 days of incubation, the bioaugmentation treatment resulted in a significant reduction in COD, 88%, 91%, and 92% in fish wastewater augmented with either Marinirhabdus sp., Marinobacter hydrocarbonoclasticus respectively, or a consortium of the two halobacteria compared with the control (non-bioaugmented) treatment (77% removal). In tall bioaugmentation treatments (79-88%) TN removal was also significantly greater than the control treatment (57%). After 9 days of incubation, the COD and TN in bioaugmentation reached the European Union's (EU) wastewater discharge standard (Level B, COD < 120 mg L-1, TN < 70 mg L-1). The addition of monoculture was effective in enhancing the removal of COD, while co-culture significantly improved TN removal. Results of 16S rDNA sequence analysis investigating the survival of these introduced bacteria showed that only Marinobacter hydrocarbonoclasticus was detected at the end of the treatment, constituting 36% of the total bacterial population when added alone to the wastewater. This study confirms the effectiveness of bioaugmentation in removing COD and TN in saline fish wastewater. The ability of Marinobacter hydrocarbonclasticus to enhance the treatment and dominate the bacterial community suggests the commercial potential of this organism for bioaugmentation of aquaculture wastewater without the need for further bioaugmentation.

Dynamic Effect of Operational Regulation on the Mesophilic BioMethanation of Grape Marc.

Wine production annually generates an estimated 11 million metric tonnes of grape marc (GM) worldwide. The diversion of this organic waste away from landfill and towards its use in the generation of renewable energy has been investigated. This study aimed to evaluate the effectiveness of operational parameters relating to the treatment regime and inoculum source in the extraction of methane from GM under unmixed anaerobic conditions at 35 °C. The study entailed the recirculation of a previously acclimated sludge (120 days) as downstream inoculum, an increased loading volume (1.3 kg) and a low substrate-to-inoculum ratio (10:3 SIR). The results showed that an incorporation of accessible operational controls can effectively enhance cumulative methane yield (0.145 m3 CH4 kg-1 VS), corresponding to higher amounts of digestible organics converted. The calculated average volumetric methane productivity equalled 0.8802 L CH4 LWork-1 d-1 over 33.6 days whilst moderate pollutant removal (43.50% COD removal efficiency) was achieved. Molecular analyses identified Firmicutes and Bacteroidetes phyla as core organisms for hydrolytic and fermentative stages in trophic relationships with terminal electron acceptors from the methane-producing Methanosarcina genus. Economic projections established that the cost-effective operational enhancements were sustainable for valorisation from grape marc by existing wineries and distilleries.

Application of Co-Culture Technology to Enhance Protease Production by Two Halophilic Bacteria, Marinirhabdus sp. and Marinobacter hydrocarbonoclasticus.

Although axenic microbial cultures form the basis of many large successful industrial biotechnologies, the production of single commercial microbial strains for use in large environmental biotechnologies such as wastewater treatment has proved less successful. This study aimed to evaluate the potential of the co-culture of two halophilic bacteria, Marinirhabdus sp. and Marinobacter hydrocarbonoclasticus for enhanced protease activity. The co-culture was significantly more productive than monoculture (1.6-2.0 times more growth), with Marinobacter hydrocarbonoclasticus being predominant (64%). In terms of protease activity, enhanced total activity (1.8-2.4 times) was observed in the co-culture. Importantly, protease activity in the co-culture was found to remain active over a much broader range of environmental conditions (temperature 25 °C to 60 °C, pH 4-12, and 10-30% salinity, respectively). This study confirms that the co-culturing of halophilic bacteria represents an economical approach as it resulted in both increased biomass and protease production, the latter which showed activity over arange of environmental conditions.

The Impacts of Different Biological Treatments on the Transformation of Explosives Waste Contaminated Sludge.

The dinitrotoluene isomers 2,4 and 2,6-dinitrotoluene (DNT) represent highly toxic, mutagenic, and carcinogenic compounds used in explosive manufacturing and in commercial production of polyurethane foam. Bioremediation, the use of microbes to degrade residual DNT in industry wastewaters, represents a promising, low cost and environmentally friendly alternative technology to landfilling. In the present study, the effect of different bioremediation strategies on the degradation of DNT in a microcosm-based study was evaluated. Biostimulation of the indigenous microbial community with sulphur phosphate (2.3 g/kg sludge) enhanced DNT transformation (82% transformation, from 300 g/L at Day 0 to 55 g/L in week 6) compared to natural attenuation over the same period at 25 °C. The indigenous microbial activity was found to be capable of transforming the contaminant, with around 70% transformation of DNT occurring over the microcosm study. 16S rDNA sequence analysis revealed that while the original bacterial community was dominated by Gammaproteobacteria (30%), the addition of sulphur phosphate significantly increased the abundance of Betaproteobacteria by the end of the biostimulation treatment, with the bacterial community dominated by Burkholderia (46%) followed by Rhodanobacter, Acidovorax and Pseudomonas. In summary, the results suggest biostimulation as a treatment choice for the remediation of dinitrotoluenes and explosives waste.

Co-Digestion of Grape Marc and Cheese Whey at High Total Solids Holds Potential for Sustained Bioenergy Generation.

At the end of fermentation, wine contains approximately 20% (w/v) of solid material, known as grape marc (GM), produced at a yield of 2 t/ha. Cheese manufacture produces cheese whey (CW), which is over 80% of the processed milk, per unit volume. Both waste types represent an important fraction of the organic waste being disposed of by the wine and dairy industries. The objective of this study was to investigate the bioenergy potential through anaerobic codigestion of these waste streams. The best bioenergy profile was obtained from the digestion setups of mixing ratio 3/1 GM/CW (wet weight/wet weight). At this ratio, the inhibitory salinity of CW was sufficiently diluted, resulting in 23.73% conversion of the organic material to methane. On average, 64 days of steady bioenergy productivity was achieved, reaching a maximum of 85 ± 0.4% CH4 purity with a maximum cumulative methane yield of 24.4 ± 0.11 L CH4 kg-1 VS. During the fermentation there was 18.63% CODt removal, 21.18% reduction of conductivity whilst salinity rose by 36.19%. It can be concluded that wine and dairy industries could utilise these waste streams for enhanced treatment and energy recovery, thereby developing a circular economy.

Elucidation of the microbial diversity in rivers in south-west Victoria, Australia impacted by rural agricultural contamination (dairy farming).

We assessed the water quality of south-west Victorian rivers impacted by the dairy industry using traditional water quality assessment together with culture-dependent (colilert/enterolert) and also culture-independent (next generation sequencing) microbial methods. The aim of the study was to identify relationships/associations between dairy farming intensity and water contamination. Water samples with high total and faecal coliforms (>1000 MPN cfu/100 ml), and with high nitrogen levels (TN) were observed in zones with a high proportion of dairy farming. Members of the genus Nitrospira, Rhodobacter and Rhodoplanes were predominant in such high cattle density zones. Samples from sites in zones with lower dairy farming activities registered faecal coliform numbers within the permissible limits (<1000 MPN cfu/100 ml) and showed the presence of a wide variety of microorganisms. However, no bacterial pathogens were found in the river waters regardless of the proportion of cattle. The data suggests that using the spatially weighted proportion of land used for dairy farming is a useful way to target at-risk sub-catchments across south west Victoria; further work is required to confirm that this approach is applicable in other regions.

Influence of bioaugmentation and biostimulation on PAH degradation in aged contaminated soils: Response and dynamics of the bacterial community.

Polycyclic aromatic hydrocarbons (PAHs) represent a group of hazardous compounds that are ubiquitous and persistent. The main aim of this study was to investigate the degradation of PAHs in chronically contaminated, aged and weathered soils obtained from a former gas plant of Australia. Biostimulation and bioaugmentation using individual isolates (Rhodococcus sp. (NH2), Achromobacter sp. (NH13), Oerskovia paurometabola (NH11), Pantoea sp. (NH15), Sejongia sp. (NH20), Microbacterium maritypicum (NH30) and Arthrobacter equi (NH21)) and a consortium of these isolates were tested during mesocosm studies. A significant reduction (99%) in PAH concentration was observed in all the treatments. In terms of the abundance of PAH-degrading genes and microbial community structure during PAH degradation, qPCR results revealed that Gram-positive bacteria were dominant over other bacterial communities in all the treatments. 16S sequencing results revealed that the inoculated organisms did not establish themselves during the treatment. However, substantial bacterial community changes during the treatments were observed, suggesting that the natural community exhibited sufficient resilience and diversity to enable an active, but changing degrading community at all stages of the degradation process. Consequently, biostimulation is proposed as the best strategy to remediate PAHs in aged, weathered and chronically contaminated soils.

Petroleum Hydrocarbon Contamination in Terrestrial Ecosystems-Fate and Microbial Responses.

Petroleum hydrocarbons represent the most frequent environmental contaminant. The introduction of petroleum hydrocarbons into a pristine environment immediately changes the nature of that environment, resulting in reduced ecosystem functionality. Natural attenuation represents the single, most important biological process which removes petroleum hydrocarbons from the environment. It is a process where microorganisms present at the site degrade the organic contaminants without the input of external bioremediation enhancers (i.e., electron donors, electron acceptors, other microorganisms or nutrients). So successful is this natural attenuation process that in environmental biotechnology, bioremediation has developed steadily over the past 50 years based on this natural biodegradation process. Bioremediation is recognized as the most environmentally friendly remediation approach for the removal of petroleum hydrocarbons from an environment as it does not require intensive chemical, mechanical, and costly interventions. However, it is under-utilized as a commercial remediation strategy due to incomplete hydrocarbon catabolism and lengthy remediation times when compared with rival technologies. This review aims to describe the fate of petroleum hydrocarbons in the environment and discuss their interactions with abiotic and biotic components of the environment under both aerobic and anaerobic conditions. Furthermore, the mechanisms for dealing with petroleum hydrocarbon contamination in the environment will be examined. When petroleum hydrocarbons contaminate land, they start to interact with its surrounding, including physical (dispersion), physiochemical (evaporation, dissolution, sorption), chemical (photo-oxidation, auto-oxidation), and biological (plant and microbial catabolism of hydrocarbons) interactions. As microorganism (including bacteria and fungi) play an important role in the degradation of petroleum hydrocarbons, investigations into the microbial communities within contaminated soils is essential for any bioremediation project. This review highlights the fate of petroleum hydrocarbons in tertial environments, as well as the contributions of different microbial consortia for optimum petroleum hydrocarbon bioremediation potential. The impact of high-throughput metagenomic sequencing in determining the underlying degradation mechanisms is also discussed. This knowledge will aid the development of more efficient, cost-effective commercial bioremediation technologies.

Viability determination of Ascaris ova in raw wastewater: a comparative evaluation of culture-based, BacLight Live/Dead staining and PMA-qPCR methods.

Accurate evaluation of viable Ascaris ova in wastewater is the key to mitigating Ascaris reinfections in endemic regions. In this study, the viability of Ascaris ova in raw wastewater was determined using three different detection methods: culture-based, BacLight Live/Dead staining and propidium monoazide-quantitative polymerase chain reaction (PMA-qPCR). Furthermore, comparative assessment of viability utilising the aforementioned detection methods was performed using seeded experiments in wastewater. The percentage of viability was: culture-based (82%), BacLight Live/Dead staining (87%) and PMA-qPCR (85%) respectively. Despite the fact that no statistical difference was shown in the viability determination among the three methods, PMA-qPCR-based viability determination would be preferable over the other two methods for evaluating potential public health risks with A. suum ova due to its accuracy, being least subjective and its rapid reaction time.

Microbial Degradation of Phenanthrene in Pristine and Contaminated Sandy Soils.

Phenanthrene mineralisation studies in both pristine and contaminated sandy soils were undertaken through detailed assessment of the activity and diversity of the microbial community. Stable isotope probing (SIP) was used to assess and identify active 13C-labelled phenanthrene degraders. Baseline profiling indicated that there was little difference in fungal diversity but a significant difference in bacterial diversity dependent on contamination history. Identification of dominant fungal and bacterial species highlighted the presence of organisms capable of degrading various petroleum-based compounds together with other anthropogenic compounds, regardless of contamination history. Community response following a simulated contamination event (14C-phenanthrene) showed that the microbial community in deep pristine and shallow contaminated soils adapted most to the presence of phenanthrene. The similarity in microbial community structure of well-adapted soils demonstrated that a highly adaptable fungal community in these soils enabled a rapid response to the introduction of a contaminant. Ten fungal and 15 bacterial species were identified as active degraders of phenanthrene. The fungal degraders were dominated by the phylum Basidiomycota including the genus Crypotococcus, Cladosporium and Tremellales. Bacterial degraders included the genera Alcanivorax, Marinobacter and Enterococcus. There was little synergy between dominant baseline microbes, predicted degraders and those that were determined to be actually degrading the contaminant. Overall, assessment of baseline microbial community in contaminated soils provides useful information; however, additional laboratory assessment of the microbial community's ability to degrade pollutants allows for better prediction of the bioremediation potential of a soil.

Large scale treatment of total petroleum-hydrocarbon contaminated groundwater using bioaugmentation.

Bioaugmentation or the addition of microbes to contaminated sites has been widely used to treat contaminated soil or water; however this approach is often limited to laboratory based studies. In the present study, large scale bioaugmentation has been applied to total petroleum hydrocarbons (TPH)-contaminated groundwater at a petroleum facility. Initial TPH concentrations of 1564 mg L-1 in the field were reduced to 89 mg L-1 over 32 days. This reduction was accompanied by improved ecotoxicity, as shown by Brassica rapa germination numbers that increased from 52 at day 0 to 82% by the end of the treatment. Metagenomic analysis indicated that there was a shift in the microbial community when compared to the beginning of the treatment. The microbial community was dominated by Proteobacteria and Bacteroidetes from day 0 to day 32, although differences at the genus level were observed. The predominant genera at the beginning of the treatment (day 0 just after inoculation) were Cloacibacterium, Sediminibacterium and Brevundimonas while at the end of the treatment members of Flavobacterium dominated, reaching almost half the population (41%), followed by Pseudomonas (6%) and Limnobacter (5.8%). To the author's knowledge, this is among the first studies to report the successful large scale biodegradation of TPH-contaminated groundwater (18,000 L per treatment session) at an offshore petrochemical facility.

Bioremediation potential of diesel-contaminated Libyan soil.

Bioremediation is a broadly applied environmentally friendly and economical treatment for the clean-up of sites contaminated by petroleum hydrocarbons. However, the application of this technology to contaminated soil in Libya has not been fully exploited. In this study, the efficacy of different bioremediation processes (necrophytoremediation using pea straw, bioaugmentation and a combination of both treatments) together with natural attenuation were assessed in diesel contaminated Libyan soils. The addition of pea straw was found to be the best bioremediation treatment for cleaning up diesel contaminated Libyan soil after 12 weeks. The greatest TPH degradation, 96.1% (18,239.6mgkg(-1)) and 95% (17,991.14mgkg(-1)) were obtained when the soil was amended with pea straw alone and in combination with a hydrocarbonoclastic consortium respectively. In contrast, natural attenuation resulted in a significantly lower TPH reduction of 76% (14,444.5mgkg(-1)). The presence of pea straw also led to a significant increased recovery of hydrocarbon degraders; 5.7log CFU g(-1) dry soil, compared to 4.4log CFUg(-1) dry soil for the untreated (natural attenuation) soil. DGGE and Illumina 16S metagenomic analyses confirm shifts in bacterial communities compared with original soil after 12 weeks incubation. In addition, metagenomic analysis showed that original soil contained hydrocarbon degraders (e.g. Pseudoxanthomonas spp. and Alcanivorax spp.). However, they require a biostimulant (in this case pea straw) to become active. This study is the first to report successful oil bioremediation with pea straw in Libya. It demonstrates the effectiveness of pea straw in enhancing bioremediation of the diesel-contaminated Libyan soil.

Biosurfactant from red ash trees enhances the bioremediation of PAH contaminated soil at a former gasworks site.

Polycyclic aromatic hydrocarbons (PAHs) are persistent contaminants that accumulate in soil, sludge and on vegetation and are produced through activities such as coal burning, wood combustion and in the use of transport vehicles. Naturally occurring surfactants have been known to enhance PAH-removal from soil by improving PAH solubilization thereby increasing PAH-microbe interactions. The aim of this research was to determine if a biosurfactant derived from the leaves of the Australian red ash (Alphitonia excelsa) would enhance bioremediation of a heavily PAH-contaminated soil and to determine how the microbial community was affected. Results of GC-MS analysis show that the extracted biosurfactant was significantly more efficient than the control in regards to the degradation of total 16 US EPA priority PAHs (78.7% degradation compared to 62.0%) and total petroleum hydrocarbons (TPH) (92.9% degradation compared to 44.3%). Furthermore the quantification of bacterial genes by qPCR analysis showed that there was an increase in the number of gene copies associated with Gram positive PAH-degrading bacteria. The results suggest a commercial potential for the use of the Australian red ash tree as a source of biosurfactant for use in the accelerated degradation of hydrocarbons.

Rhizoremediation of phenanthrene and pyrene contaminated soil using wheat.

Rhizoremediation, the use of the plant rhizosphere and associated microorganisms represents a promising method for the clean up of soils contaminated with polycyclic aromatic hydrocarbons (PAHs) including phenanthrene and pyrene, two model PAHs. Although numerous studies have been published reporting the degradation of phenanthrene and pyrene, very few evaluate the microbial basis of the rhizoremediation process through the application of molecular tools. The aim of this study was to investigate the effect of wheat on the degradation of two model PAHs (alone or in combination) and also on soil bacterial, fungal and nidA gene (i.e. a key gene in the degradation of pyrene) communities. The addition of wheat plants led to a significant enhancement in the degradation of both phenanthrene and pyrene. In pyrene-contaminated soils, the degradation rate increased from 15% (65 mg/kg) and 18% (90 mg/kg) in unplanted soils to 65% (280 mg/kg) and 70% (350 mg/kg) in planted treatments while phenanthrene reduction was enhanced from 97% (394 mg/kg) and 87% (392 mg/kg) for unplanted soils to 100% (406 mg/kg) and 98% (441 mg/kg) in the presence of wheat. PCR-DGGE results showed that the plant root let to some changes in the bacterial and fungal communities; these variations did not reflect any change in hydrocarbon-degrading communities. However, plate counting, traditional MPN and MPN-qPCR of nidA gene revealed that the wheat rhizosphere led to an increase in the total microbial abundance including PAH degrading organisms and these increased activities resulted in enhanced degradation of phenanthrene and pyrene. This clearer insight into the mechanisms underpinning PAH degradation will enable better application of this environmentally friendly technique.

Necrophytoremediation of phenanthrene and pyrene in contaminated soil.

In this study, the effect of necrophytoremediation, using pea and wheat straws on the remediation soil contaminated with two common polycyclic aromatic hydrocarbons (PAHs), phenanthrene and pyrene alone or in combination was investigated. In addition, monitoring of the population of PAH-utilising microorganisms together with PCR-Denaturing Gradient Gel Electrophoresis (DGGE)-sequencing methods were used to further elucidate the effect of straw addition on the bacterial, fungal and nidA gene (a functional gene involved in the degradation of PAHs) communities. The addition of pea straw had a positive effect on the degradation of PAHs, especially for pyrene. For example, the addition of pea straw to pyrene-contaminated soil resulted in an increase in the degradation of pyrene from 15% (66 mg kg(-1)) in the corresponding control to 70% (301 mg kg(-1)). The results from the most probable number (MPN) of PAH-utilising microorganisms and PCR-DGGE-sequencing methods indicated that the addition of straw led to an increase in microbial hydrocarbonoclastic biomass rather than changes in microbial diversity. For example, in pyrene-contaminated soil, the number of PAH-utilising microorganisms in the soil amended with pea straw reached 5.6 log10 CFU g(-1) dry soil, ~13-fold increase when compared with the numbers present in the control soil (4.5 log10 CFU g(-1) dry soils); however, the Shannon diversity index did not increase significantly. The sequencing of bands of interest from DGGE profiles revealed the presence of Actinobacteria, Firmicutes and Proteobacteria in the bacterial community. For fungi, sequenced bands belonged to Ascomycota, Basidiomycota and Mucoromycotina. In summary, this study has shown that necrophytoremediation using pea straw represents a promising biostimulation and cost effective agent which can be used for the bioremediation of PAH-contaminated soils.

Substrate-to-inoculum ratio drives solid-state anaerobic digestion of unamended grape marc and cheese whey.

Inoculation dose is a key operational parameter for the solid-state anaerobic digestion (SS-AD) of lignocellulosic biomass, maximum methane recovery, and stable digester performance. The novelty of this study was the co-digestion of unamended full-strength grape marc and cheese whey for peak methane extraction at variable inoculation levels. An acclimatised digestate from a preceding anaerobic treatment was used as a downstream inoculum. The impact of inoculum size (wet weight) was evaluated at 0/10, 5/5, 7/3 and 9/1 substrate-to-inoculum (S/I) ratios, corresponding to an initial concentration of 20-30% total solids (TS) in digesters over 58 days at 45°C. The optimal 7/3 S/I produced the highest cumulative methane yield, 6.45 L CH4 kg-1 VS, coinciding with the lowest initial salinity at 11%; the highest volumetric methane productivity rate of 0.289±0.044 L CH4 LWork-1 d-1; the highest average COD/N ratio of 9.88; the highest final pH of 9.13, and a maximum 15.07% elemental carbon removal; for a lag time of 9.4 days. This study identified an optimal inoculation dose and opens up an avenue for the direct co-digestion of grape marc and cheese whey without requirements for substrate pretreatment, thus improving the overall bioenergy profile of the winery and dairy joint resource recovery operations.

Phylogenetic diversity of fungal communities in areas accessible and not accessible to tourists in Naracoorte Caves.

The fungal diversity in areas accessible and not accessible to tourists at UNESCO World Heritage-listed Naracoorte Caves was investigated with culture-dependent and culture-independent techniques for assistance in cave management protocol development. The caves were selected based on tourist numbers and configurations: Stick Tomato (open, high numbers), Alexandra (lockable openings, high numbers) and Strawhaven (control; no access). Culture-based survey revealed Ascomycota dominance irrespective of sampling area with Microascales (Trichurus sp.) being most frequently isolated. Some Hypocreales-like sequences belonging to Fusarium sp., Trichoderma sp. and Neonectria sp. (Stick Tomato) were cultured only from areas not accessible to tourists. These orders also were detected by DGGE assay irrespective of sampling area. The predominance of Ascomycota (especially Microascales) suggested their important ecological roles in these caves. Culture-independent analysis showed higher Shannon fungal diversity values (from ITS-based DGGE profiles) in tourist-accessible areas of these caves than in inaccessible areas with the fungal community banding patterns being substantially different in Stick Tomato Cave. Further investigations are needed to determine the cause of the differences in the fungal communities of Stick Tomato Cave, although cave-related factors such as use, configuration and sediment heterogeneity might have contributed to these differences.

The effects of vehicular emissions on the activity and diversity of the roadside soil microbial community.

Motor vehicles emit a variety of pollutants including metals, petroleum hydrocarbons and polycyclic aromatic hydrocarbons (PAHs). The relationships between metals, petroleum hydrocarbons and PAHs, soil respiration and microbial diversity (fungi and bacteria) were studied using control (n = 3) and roadside soils (n = 27) with different exposure periods to vehicle emissions (2-63 years). Bacterial diversity was found to be higher than control sites (P = 0.002) but was the same across different categories of road age (P = 0.328). Significant (r = -0.49, P = 0.007) contrasting behaviour of fungal and bacterial diversity was reported, with diversity increasing across all road types for bacteria and decreasing across all road types for fungi compared to control soils. Analysis of the bacterial community identified three distinct clusters, separated on age of contamination, suggesting that roadside bacterial communities change over time with pollution from vehicles with the potential development of metal resistant bacteria in roadside soils. In contrast, for fungal communities, a reduction in diversity with time of exposure to roadside vehicle emissions was observed suggesting the potential for reduced ecosystem functionality and soil health in roadside soils. This is the first study in the published literature to include both bacterial and fungal responses from aged roadside soils. The results from this study suggest that normal functionality of soil ecosystem services is being affected in roadside soils, potentially globally.

Can biochar be an effective and reliable biostimulating agent for the remediation of hydrocarbon-contaminated soils?

Petroleum hydrocarbons represent one of the most common soil contaminants, whose presence poses a significant risk to soil biota and human health; for example, in Europe, hydrocarbon contamination accounts for more than 30% of contaminated sites. The use of biochar as a proposed alternative to the conventional remediation of soil contaminated with petroleum hydrocarbons has gained credence in recent times because of its cost-effectiveness and environmentally friendly nature. Biochar is a carbonaceous material produced by heating biomass in an oxygen-limited environment at high temperature. This review provides an overview of the application of biochar to remediate petroleum hydrocarbon-contaminated soils, with emphasis on the possibility of biochar functioning as a biostimulation agent. The properties of biochar were also examined. Furthermore, the mechanism, ecotoxicological impact and possible factors affecting biochar-based remediation are discussed. The review concludes by examining the drawbacks of biochar use in the remediation of hydrocarbon-contaminated soils and how to mitigate them. Biochar impacts soil microbes, which may result in the promotion of the degradation of petroleum hydrocarbons in the soil. Linear regression between bacterial population and degradation efficiency showed that R2 was higher (0.50) and significant in treatment amended with biochar or both biochar and nutrient/fertiliser (p < 0.01), compared to treatment with nutrient/fertiliser only or no amendment (R2 = 0.11). This suggest that one of the key impacts of biochar is enhancing microbial biomass and thus the biodegradation of petroleum hydrocarbons. Biochar represents a promising biostimulation agent for the remediation of hydrocarbon-contaminated soil. However, there remains key questions to be answered.