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- Acinetobacter rudis sp. nov., isolated from raw milk and raw wastewaterPublication . Vaz-Moreira, Ivone; Novo, Ana; Hantsis-Zacharov, Elionora; Lopes, Ana Rita; Gomila, Margarita; Nunes, Olga C.; Manaia, Célia M.; Halpern, MalkaTwo bacterial strains, G30T and A1PC16, isolated respectively from raw milk and raw wastewater, were characterized using a polyphasic approach. Chemotaxonomic characterization supported the inclusion of these strains in the genus Acinetobacter, with Q-8 and Q-9 as the major respiratory quinones, genomic DNA G+C contents within the range observed for this genus(38–47 mol%) and C16 : 0, C18 : 1v9c and C16 : 1v7c/iso-C15 : 0 2-OH as the predominant fatty acids. The observation of 16S rRNA gene sequence similarity lower than 97% with other Acinetobacter species with validly published names led to the hypothesis that these isolates could represent a novel species. This hypothesis was supported by comparative analysis of partial sequences of the genes rpoB and gyrB, which showed that strains G30T and A1PC16 did not cluster with any species with validly published names, forming a distinct lineage. DNA–DNA hybridizations confirmed that the two strains were members of the same species, which could be distinguished from their congeners by several phenotypic characteristics. On the basis of these arguments, it is proposed that strains G30T and A1PC16 represent a novel species, for which the name Acinetobacter rudis sp. nov. is proposed. The type strain is strain G30T (5LMG 26107T 5CCUG 57889T 5DSM 24031T 5CECT 7818T).
- Paenibacillus residui sp. nov., isolated from urban waste compostPublication . Vaz-Moreira, Ivone; Figueira, Vânia; Lopes, Ana Rita; Pukall, Rüdiger; Spröer, Cathrin; Schumann, Peter; Nunes, Olga C.; Manaia, Célia M.Two bacterial strains, MC-246T and MC-247, were isolated from municipal urban waste compost and characterized by a polyphasic approach. Both isolates were Gram-stain-variable, endosporeforming rods that were catalase-, oxidase- and b-galactosidase-positive, and able to grow at 25– 50 6C and pH 7.0–9.0, with optimum growth at 37 6C and pH 7. The predominant cellular fatty acids were anteiso-C15 : 0, iso-C15 : 0, iso-C16 : 0, anteiso-C17 : 0 and iso-C17 : 0; the major respiratory quinone was menaquinone MK-7; the cell wall peptidoglycan was of type A1c; and the DNA G+C content was 49 mol%. These characteristics, as well as data from 16S RNA gene sequence analysis, showed that these strains were affiliated with the genus Paenibacillus; the type strains of Paenibacillus ginsengarvi and Paenibacillus hodogayensis were among their closest neighbours (,94.2 % sequence similarity). Nevertheless, the hypothesis that strains MC246T and MC-247 could represent a novel species was supported by the low 16S rRNA gene sequence similarity values shared with other members of the genus Paenibacillus and by the observation of distinct biochemical and physiological traits. Strains MC-246T and MC-247 shared 99.6 % 16S rRNA gene sequence similarity and showed almost identical MALDI-TOF mass spectra, but could be distinguished at the phenotypic and genotypic level. However, DNA–DNA hybridization between strains MC-246T and MC-247 resulted in values above 70 % indicating that both organisms represent a single species, for which the name Paenibacillus residui sp. nov. is proposed; the type strain is MC-246T (5DSM 22072T 5CCUG 57263T).
- Removal of microorganisms and antibiotic resistance genes from treated urban wastewater: a comparison between aluminium sulphate and tannin coagulantsPublication . Grehs, Bárbara W. N.; Lopes, Ana Rita; Moreira, Nuno F. F.; Fernandes, Telma; Linton, Maria A. O.; Silva, Adrián M. T.; Manaia, Célia M.; Carissimi, Elvis; Nunes, Olga C.The presence of antibiotic resistant-bacteria (ARB) and antibiotic resistance genes (ARG) in treated effluents of urban wastewater treatment plants (WWTP) may represent a threat to the environment and public health. Therefore, cost-effective technologies contributing to minimize loads of these contaminants in the final effluents of WWTP are required. This study aimed at assessing the capacity of coagulation to reduce the ARB&ARG load in secondary treated urban wastewater (STWW), as well as the impact of the process on the structure and diversity of the bacterial community. Coagulation performance using aluminium sulphate, a synthetic substance, and tannins, a biowaste, was compared. Samples were analysed immediately before (STWW) and after the coagulation treatment (Alu, Tan), as well as after 3-days storage in the dark at room temperature (RSTWW, RAlu, RTan), to assess possible reactivation events. Both coagulants decreased the turbidity and colour and reduced the bacterial load (16S rRNA gene copy number, total heterotrophs (HET), and ARB (faecal coliforms resistant to amoxicillin (FC/AMX) or ciprofloxacin (FC/CIP) up to 1–2 log immediately after the treatment. Both coagulants reduced the load of intl1, but in average, aluminium sulphate was able to decrease the content of the analysed ARGs (blaTEM and qnrS) to lower levels than tannin. Reactivation after storage was observed mainly in RTan. In these samples the load of the culturable populations and qnrS gene prevalence increased, sometimes to values higher than those found in the initial wastewater. Reactivation was also characterized by an increment in Gammaproteobacteria relative abundance in the bacterial community, although with distinct patterns for RTan and RAlu. Curvibacter, Undibacterium and Aquaspirillum were among the most abundant genera in RAlu and Aeromonas, Pseudomonas and Stenotrophomonas in RTan. These bacterial community shifts were in agreement with the variations in the culturable bacterial counts of HET for RTan and FC/CIP for RAlu. In summary, the overall performance of aluminium sulphate was better than that of tannins in the treatment of treated urban wastewater.
- Wastewater reuse in irrigation: a microbiological perspective on implications in soil fertility and human and environmental healthPublication . Becerra-Castro, Cristina; Lopes, Ana Rita; Vaz-Moreira, Ivone; Silva, Elisabete F.; Manaia, Célia M.; Nunes, Olga C.The reuse of treated wastewater, in particular for irrigation, is an increasingly common practice, encouraged by governments and official entities worldwide. Irrigation with wastewater may have implications at two different levels: alter the physicochemical and microbiological properties of the soil and/or introduce and contribute to the accumulation of chemical and biological contaminants in soil. The first may affect soil productivity and fertility; the second may pose serious risks to the human and environmental health. The sustainable wastewater reuse in agriculture should prevent both types of effects, requiring a holistic and integrated risk assessment. In this article we critically review possible effects of irrigation with treated wastewater, with special emphasis on soil microbiota. The maintenance of a rich and diversified autochthonous soil microbiota and the use of treated wastewater with minimal levels of potential soil contaminants are proposed as sine qua non conditions to achieve a sustainable wastewater reuse for irrigation.
- Comparative study of the microbial diversity of bulk paddy soil of two rice fields subjected to organic and conventional farmingPublication . Lopes, Ana R.; Faria, Cátia; Prieto-Fernández, Ángeles; Trasar-Cepeda, Carmen; Manaia, Célia M.; Nunes, Olga C.Two adjacent paddies of an experimental rice field, subjected to organic and conventional farming, were characterized aiming the comparative assessment of microbiological variations occurring in the bulk paddy soil over the rice cycle. This study comprehended the simultaneous characterization of general physicochemical soil properties [total carbon and nitrogen, pH (H2O and KCl), C:N ratio and water content], biochemical properties [enzymatic activities and Community Level Physiological Profiles (CLPP)], the estimation of cultivable organisms (enumeration of fast growing heterotrophic bacteria, actinomycetes and fungi) and the assessment of bacterial diversity using a culture-independent method (PCR-DGGE fingerprinting). The linkage of the parameters measured was analysed by canonical correspondence analysis (CCA). CCA ordination plots of the CLPP showed a similar pattern of microbial functional activity in both agronomic management systems, except in June. Enzymatic activity, water content and fungi counts were the main factors affecting the observed CLPP time variation. Such a variation was not expressed by the Shannon and evenness indices, which did not evidence significant differences in the bacterial and functional diversity between or within farming type over the analysed period. The cluster and CCA analyses of the DGGE profiles allowed the distinction of the bacterial communities of both paddies, with temporal variations being observed in the organically managed field but not in the conventional paddy. Enzymatic activity, pH and molinate content were the factors which most contributed to the observed variations. Altogether these results underline the functional redundancy of the rice paddy soil and evidence the temporal variations on the metabolic activity of soil, irrespective of farming type.
- Shinella fusca sp. nov., isolated from domestic waste compostPublication . Vaz-Moreira, Ivone; Faria, Cátia; Lopes, Ana R.; Svensson, Liselott A.; Moore, Edward R. B.; Nunes, Olga C.; Manaia, Célia M.A bacterium, designated strain DC-196T, isolated from kitchen refuse compost was analysed by using a polyphasic approach. Strain DC-196T was characterized as a Gram-negative short rod that was catalase- and oxidase-positive, and able to grow at 10–40 6C, pH 6–9 and in NaCl concentrations as high as 3%. Chemotaxonomically, C18 : 1 was observed to be the predominant cellular fatty acid and ubiquinone 10 (Q10) was the predominant respiratory quinone. The G+C content of the genomic DNA was determined to be 66 mol%. On the basis of the genotypic, phenotypic and chemotaxonomic characteristics, strain DC-196T was assigned to the genus Shinella, although with distinctive features. At the time of writing, 16S rRNA gene sequence similarities of 97.6–96.8% and the low DNA–DNA hybridization values of 38.2–32.2% with the type strains of the three recognized Shinella species confirmed that strain DC-196T represents a novel species of the genus, for which the name Shinella fusca sp. nov. is proposed (type strain DC-196T5CCUG 55808T5LMG 24714T).
- Characterization of bacterial communities from Masseiras, a unique Portuguese greenhouse agricultural systemPublication . Becerra-Castro, Cristina; Lopes, Ana Rita; Teixeira, Sara; Silva, M. Elisabete F.; Pimenta, Elisabete; Manaia, Célia M.; Nunes, Olga C."Masseiras" is an ancient Portuguese agriculture system, where soil was developed from sand dunes enriched with seaweeds over more than a century. Due to the importance for the local economy, this system evolved for greenhouse structures. In this study we compared the bacterial community composition and structure of "Masseiras" soil, aiming at assessing the potential impact of different agricultural practices. The bulk soil of two greenhouses (following or not the recommended agriculture good practices, FGP and NFGP, respectively) was compared based on their physicochemical properties and bacterial community. In both FGP and NFGP, Proteobacteria, Acidobacteria, Actinobacteria, Bacteroidetes, Firmicutes, and Gemmatimonadetes were in a proportion of 5:1:1:1:1:1. However, the bacterial community of soil FGP was richer and more diverse than that of soil NFGP. Members of the classes Bacilli and Gemm-1, with higher relative abundance in NFGP and FGP, respectively, were those contributing most for distinguishing the bacterial communities of both soils. The differences in the structure of the bacterial communities correlated (Mantel test) with some soil physicochemical properties, such as electrical conductivity and nitrate and Zn contents, which were significantly higher in soil NFGP than in soil FGP.
- Relationships among bulk soil physicochemical, biochemical, and microbiological parameters in an organic alfalfa-rice rotation systemPublication . Lopes, Ana R.; Bello, Diana; Prieto-Fernández, Ángeles; Trasar-Cepeda, Carmen; Manaia, Célia M.; Nunes, Olga C.The microbial communities of bulk soil of rice paddy fields under an ancient organic agriculture regimen, consisting on an alfalfa-rice rotation system, were characterized. The drained soil of two adjacent paddies at different stages of the rotation was compared before rice seeding and after harvesting. The relationships among the soil microbial, physicochemical, and biochemical parameters were investigated using multivariate analyses. In the first year of rice cropping, aerobic cultivable heterotrophic populations correlated with lineages of presumably aerobic bacteria (e.g., Sphingobacteriales, Sphingomonadales). In the second year of rice cropping, the total C content correlated with presumable anaerobic bacteria (e.g., Anaerolineae). Independently of the year of rice cropping, before rice seeding, proteolytic activity correlated positively with the cultivable aerobic heterotrophic and ammonifier populations, the soil catabolic profile and with presumable aerobes (e.g., Sphingobacteriales, Rhizobiales) and anaerobes (e.g., Bacteroidales, Anaerolineae). After harvesting, strongest correlations were observed between cultivable diazotrophic populations and bacterial groups described as comprising N-2 fixing members (e.g., Chloroflexi-Ellin6529, Betaproteobacteria, Alphaproteobacteria). It was demonstrated that chemical parameters and microbial functions were correlated with variations on the total bacterial community composition and structure occurring during rice cropping. A better understanding of these correlations and of their implications on soil productivity may be valid contributors for sustainable agriculture practices, based on ancient processes.
- Candidimonas nitroreducens gen. nov., sp. nov. and Candidimonas humi sp. nov., isolated from sewage sludge compostPublication . Vaz-Moreira, Ivone; Figueira, Vânia; Lopes, Ana R.; De Brandt, Evie; Vandamme, Peter; Nunes, Olga C.; Manaia, Célia M.Two bacterial strains (SC-089T and SC-092T) isolated from sewage sludge compost were characterized by using a polyphasic approach. The isolates were Gram-negative short rods, catalase- and oxidase-positive, and showed good growth at 30 °C, at pH 7 and with 1 % (w/v) NaCl. Ubiquinone 8 was the major respiratory quinone, and phosphatidylethanolamine, phosphatidylglycerol and diphosphatidylglycerol were amongst the major polar lipids. On the basis of 16S rRNA gene sequence analysis, the strains were observed to be members of the family Alcaligenaceae, but could not be identified as members of any validly described genus. The low levels of 16S rRNA gene sequence similarity to other recognized taxa, together with comparative analysis of phenotypic traits and chemotaxonomic markers, supported the proposal of a new genus within the family Alcaligenaceae, for which the name Candidimonas gen. nov. is proposed. Strains SC-089T and SC-092T, which shared 99.1 % 16S rRNA gene sequence similarity, could be differentiated at the phenotypic level, and DNA–DNA hybridization results supported their identification as representing distinct species. The names proposed for these novel species are Candidimonas nitroreducens sp. nov. (type strain, SC-089T = LMG 24812T = CCUG 55806T) and Candidimonas humi sp. nov. (type strain, SC-092T = LMG 24813T = CCUG 55807T).
- Bacterial community variations in an alfalfa-rice rotation system revealed by 16S rRNA gene 454-pyrosequencingPublication . Lopes, Ana R.; Manaia, Célia M.; Nunes, Olga C.Crop rotation is a practice harmonized with the sustainable rice production. Nevertheless, the implications of this empirical practice are not well characterized, mainly in relation to the bacterial community composition and structure. In this study, the bacterial communities of two adjacent paddy fields in the 3rd and 4th year of the crop rotation cycle and of a nonseeded subplot were characterized before rice seeding and after harvesting, using 454-pyrosequencing of the 16S rRNA gene. Although the phyla Acidobacteria, Proteobacteria, Chloroflexi, Actinobacteria and Bacteroidetes predominated in all the samples, there were variations in relative abundance of these groups. Samples from the 3rd and 4th years of the crop rotation differed on the higher abundance of groups of presumable aerobic bacteria and of presumable anaerobic and acidobacterial groups, respectively. Members of the phylum Nitrospira were more abundant after rice harvest than in the previously sampled period. Rice cropping was positively correlated with the abundance of members of the orders Acidobacteriales and Solibacterales' and negatively with lineages such as Chloroflexi Ellin6529'. Studies like this contribute to understand variations occurring in the microbial communities in soils under sustainable rice production, based on real-world data.