Soil Microbiology Research Articles

The soil microbiomes of the Brazilian Cerrado

The slightly exuberant appearance suggests a misleading idea of an insignificant diversity of microorganisms in Cerrado soils. Thus, a review is necessary, gathering essential information on the microbiology of the Cerrado soil, and its biogeochemical processes, as well as the effects of anthropogenic impacts on this important biome. We reviewed studies of microbiota, bacteria, archaea, and fungi, from the Cerrado soil, which were carried out by metagenomic methods, or amplicons sequencing. Also, we evaluated the descriptions of the Cerrado’s characteristics, and describe the implications of the expansion of agricultural areas, anthropic impacts, and wildfires on the soil microbiota. The acidic soil, subject to variations in nutrient cycling depending on the climatic season, reflects on the structures of the communities of the main bacterial groups of Proteobacteria, Acidobacteria, and Actinobacteria. Members of the Archaea are also abundant in the Cerrado soil, with an emphasis on representatives of the groups Euryarchaeota, Crenarchaeota, and Thaumarchaeota. The fungi found in Cerrado soils are normally dominated by Ascomycota and Basidiomycota, which exert influence of the impact of land use, such as grazing and agriculture. Despite its immense microbial biodiversity, the Cerrado constantly suffers the impacts of the “new agricultural frontier,” which has been devastating the biome through the cultivation of sugarcane, maize, cotton, and soybean monocultures, in particular. These activities, with consequent loss of native areas of the Cerrado, modify the microbiome present in the soil, changing the flow of nutrients and interfering in biogeochemical processes.

On the approach to the analysis of research vectors as the case study of Microbiology subject area

It is very important to identify the most developing research areas. The analysis of terminology dynamics, as exemplified by microbiology, enables to make conclusions on the current changes within the corresponding branch of science. The terminological dynamics of the most cited publications demonstrates high relevance of research findings demanded by scientific community. As exemplified by our Microbiology subject index, we introduce the method of identification of the most developing research problems based on key words frequency distribution. We suggest that the share of unique topic-oriented key words in the articles which belong to the same topic-oriented group is a variable that correlates with the range of research studies, methods, and the diversity of microorganisms: the greater is the number of these words, the greater diversity of topics is comprised by scientific papers in the field. Within this study, the group of papers falling under the heading of Genetics of yeasts and microfungi is leading in the number of papers with the number of topic-oriented key words amounting to 83%. Soil microbiology, Geomicrobiology, Pathogen-host interactions, and Bacterial Genetics also belong to the most developing topics.

Microbiota in the Rhizosphere of Cereal Crops

Today, spelt wheat grain is used to produce high quality food. Intermediate wheatgrass is a promising crop for prairie restoration. One of the elements of biologization is the influence of growing crops on the microbiota of soil rhizosphere. The microbiota of spelt wheat and intermediate wheatgrass soil rhizosphere remains insufficiently studied. Aim. To study the number of individual groups of microbiota in dynamics in the rhizosphere of cereal crops (spelt wheat, intermediate wheatgrass) depending on the weather conditions and the phase of plants development. Methods. Classical microbiological, statistical methods were used in the work. In particular, the study of the number of microorganisms of different ecological and trophic groups (ammonifying, nitrifying, cellulolytic and nitrogen-fixing) was carried out according to generally accepted methods in soil microbiology. The reliability of the influence of factors was determined by the probability value «р» level which was calculated using STATISTICA 8 program. Results. The amount of ammonifying and cellulolytic microorganisms in the soil rhizosphere of spelt wheat is significantly higher compared to soft wheat. The rhizosphere microbiota amount of the intermediate wheatgrass on the 2–3 year of cultivation was more resistant to adverse environmental factors compared to soft wheat. The soil rhizosphere microbiota did not change a lot depending on the phase of plant development during the vegetation period of cereal crops (spelt wheat, intermediate wheatgrass). Conclusions. The formation of rhizosphere microbiota of spelt wheat and intermediate wheatgrass was first analyzed under the conditions of the Right-Bank forest-steppe of Ukraine. The conducted studies indicate the feasibility of growing and use of spelt wheat in breeding programs to create cultivars of soft wheat with higher activity of rhizosphere microbiota. The number of ammonifying, nitrifying and cellulolytic microorganisms of soil rhizosphere of intermediate wheatgrass was significantly higher compared to soft wheat during all growth stages. The conducted studies confirm the practical application of intermediate wheatgrass to preserve and increase soil fertility. Intermediate wheatgrass can be grown for up to three years in one field, as microbiological activity reaches its maximum development.

Influence of biofertilizers and vermicompost on soil health and Soil Microbiology

Influence of biofertilizers and vermicompost on soil health and Soil Microbiology

Maximization of Nutrient use Efficiency and Yield through Application of Biofertilizers in Field Pea (Pisum sativum L.)

Background: The negative effects of continuous use of chemical fertilizers on soil microbiology and agricultural sustainability are well established. The chemical fertilizers load in environment can be minimized by combined application of fertilizers and biofertilizers in crops like pulses which require less fertilizer-N and respond well to the use of biofertilizers. The objectives of the study were to see the effect of different biofertilizers in reduction of total fertilizer use and the response of field pea to combined application of chemical- and bio-fertilizers in terms of growth, yield and NUE.Methods: A field experiment was conducted during 2 consecutive rabi season of 2017-18 and 2018-19 at KVK, Chhatarpur, Jabalpur (Madhya Pradesh), India to evaluate the application of fertilizers and biofertilizers on growth parameter, nodulation, nutrient content and uptake, nutrient use efficiency, yield and economics of field pea. The experiment was laid out in randomized block design with five treatments in four replications. Treatments comprised of Control, Recommended dose (RD) of NPK (20: 60: 20 kg N, P2O5 and K2O/ha, respectively) (RDNPK), RDNPK + seed inoculation with Rhizobium @ 20 g/kg seed (RDNPK +R), RDNPK+R+phosphate solubilizing bacteria @ 20 g each/kg seed (RDNPK + R+PSB) and 75% of RDNPK + R+ PSB + potash solubilizing bacteria @ 5 kg/ha (75% RDNPK+ R+PSB+KSB). All other practices followed as per recommendation for the region and different observations and indices were recorded by following standard procedures.Conclusion: The application of 75% RDNPK+R+PSB+KSB was found best treatment among all others which resulted in highest grain yield (1682 kg/ha), protein content (23.1%), protein yield (388.5 kg/ha), net return (Rs. 46 623/ha) and B:C ratio (2.94). The nutrient use efficiency such as Partial Factor Productivity (PFP), Agronomic efficiency (AE), Physiological Efficiency (PE) and Economic Efficiency (EE) were also higher under combined application of fertilizers and biofertilizers. Thus, 75% RDNPK along with combined application of biofertilizers (R+PSB+KSB) may be applied for higher yield and return from field pea.

Effects of biological agents on soil microbiology, enzyme activity and fruit quality of kiwifruit with root rot

Context Root rot is common for kiwifruit (Actinidia chinensis var. diliciosa) plants. It is not easily controlled by chemical methods, and biological agents are an alternative. Aims This study investigated the effects of biological agents on the activity and diversity of soil microorganisms, soil enzyme activity and physicochemical indices during the growth period of kiwifruit plants. Methods There were four treatments: (1) root-rot plants applied with traditional fertiliser (RP); (2) root-rot plants applied with biological agents CY (CY); (3) root-rot plants applied with biological agents CL (CL); and (4) healthy plants with traditional fertiliser (HP). Most samples were taken during the growth period of kiwifruit plants, while activities of defensive-related enzymes and quality of kiwifruits were tested in the fructescence period. Key results CY and CL treatments significantly improved microbial activity, changed microbial structure, increased the diversity, richness and uniformity of microbial species, and altered the relative utilisation ratio of six carbon sources. Soil nutrients of kiwifruit plants with root rot improved with CY and CL compared to RP. Health of kiwifruit plants with CY and CL also improved compared to RP. Activities of defensive-related enzymes in CY and CL treatments were significantly higher than in RP treatment (P < 0.05). Fruit quality indices were also higher than RP. Conclusions CY and CL changed the microbial communities in soil, and improved soil nutrients and plant health. Implications By altering the soil microbial structure, biological agents used to control root rot in kiwifruit plants improved tree health and fruit quality, and provide an alternative to chemical control of root rot.

Affecting Factors of Soil Microorganisms and Their Research Prospects

There are many factors affecting the uncertainty of soil microorganisms. This paper aims at reviewing the influencing factors and research progress of soil microorganisms, such as bacteria, fungi and actinomycetes, soil microbial biomass, such as microbial biomass carbon, microbial biomass nitrogen and microbial biomass P, in the ecological distribution of Soil microbes has been discussed from the aspects of time, space and plant communities etc., focused on effects of natural factors including soil moisture, soil temperature, soil PH, soil nutrients, soil salinity, etc. and artificial measures including fertilization and spraying rare earth elements, grazing, different utilization and tillage practices, crop rotation, burning, fencing, tourism trampling, soil pollution, etc. on soil microbial. Although there are many reports about soil microbiology, overall it is still limited. And look into the distance for future research by combining with our project team for many years of work.

Intraspecies variability of the 16S rRNA gene of the soil bacteria Acinetobacter lwoffii and Paenibacillus taichungensis

The functioning of well-studied key groups of soil microorganisms depends on the microbial ecosystem in which they function in interaction with species belonging to other ecological groups. An analysis of the composition of the soil microbiota, and the possibility of analyzing the number of representatives of specific species, remains an urgent task of soil microbiology. Such analysis could be performed using modern and expensive methods such as metagenomics and mass-spectrometry, but some questions could also be answered using real-time PCR. This well-known approach is rather cheap for massive analysis and is ready to present reproducible results for practical agricultural applications. Understanding the variability of the primary structure of 16S rRNA is key to the reliable identification of bacterial species and provides an opportunity to choose the optimal pathways for their detection by PCR. In this work, analysis of the sequences of 16S rRNA of two species of soil bacteria, Acinetobacter lwoffii and Paenibacillus taichungensis is carried out. The most variable and most conservative areas on the level of species are detected. It was proved that conventional variable and conservative areas of the gene have on average nearly the same level of variability on the intraspecies level.

Traces of ancient agriculture in the soil around the archaeological sites (A case study from Northern Caucasus, Russia)

The article describes the results of research of traces of ancient and medieval agriculture in the vicinity of the settlement Podkumskoe-3 of the Alan culture (5th-8th centuries) in the Northern Caucasus, Russia. Methods of soil science, soil microbiology, archaeology, and archaeobotanical analysis were used. Soil sections were made at different distances from the settlement (up to 1200 m). It is shown that ancient and medieval agriculture left a noticeable trace in the soil in the form of ceramics, charred grains, and straw of cereals, and also led to changes in the chemical and biological properties of soils within the potential economic zone of the site. The soils within the 500-600-m zone around the settlement Podkumskoe-3 bear traces of plowing and fertilizing in the Early Middle Ages. At a greater distance from the settlement, traces of medieval agriculture disappear, but everywhere there are traces of plowing in the Late Bronze Age in the form of ceramics of the Koban culture. The most reliable indicators of ancient agriculture are, in addition to ceramics, coals and charred remains of grains of cultivated cereals and straw. However, charred grains and straw are preserved only in the lower layers of the soil, corresponding to the first stage of plowing. In the more recent layers, which were plowed for a long time, the coals are destroyed. The fertilization zone is determined by the abundance of thermophilic microorganisms. They got into the soil with manure after self-heating of animal dung during decomposing. This indicator well marks the fertilization zone in the Early Middle Ages and shows a close correlation with the ceramics of this time. In soils where only Bronze Age ceramics are present, the number of thermophilic bacteria is at the level of background soils.

Biochemical activity and microbial biomass in wetlands (Vereda) and well-drained soils under native vegetation types in Brazilian Cerrado

The Cerrado is the second largest Brazilian biome, a world biodiversity hotspot and home to important hydrographic basins. In this biome, native vegetation types play crucial environmental roles as natural repositories of carbon, water and biodiversity. However, the Cerrado is threatened by deforestation and land degradation. The soil microbiology knowledge of these areas would provide a better understanding of how this threatened environment functions and a basis for monitoring and conservation. This study investigates soil respiration, microbial biomass and enzymes activity of the native wetlands (veredas), and well-drained soils under native cerrado sensu stricto and semi-deciduous dry forest. Vereda soils produced the highest levels of soil biological attributes, while the semi-deciduous dry forest produced intermediate values and the Cerrado, the lowest values. Regardless of season, differences in soil activity were best correlated with five biological attributes: basal respiration, FDA, urease, β-glucosidase and acid phosphatase activity. Partial redundancy analysis indicated that variance in soil microbial and biochemical attributes was better correlated with soil physicochemical attributes than with vegetation type. Redundancy analysis and a Pearson correlation showed that the differentiating biological variables were best correlated with nitrogen concentration. Vereda soils have a far greater biochemical potential for metabolizing organic carbon under standard (i.e. aerobic) conditions, suggesting that draining vereda soils may lead to losses of organic matter and increased greenhouse gas emissions. The findings of this study provide a basis of ecological information on native Cerrado soils that can be used to manage and monitor the quality of this threatened biome.

A novel generalized odd log-logistic Maxwell-based regression with application to microbiology

The presence of bimodality, heteroskedasticity, zero-inflation and nonlinear effects in covariables is common in several real data applications. In this context, new regressions are proposed for data with all these characteristics. The estimation of parameters follows the maximum likelihood method. For different fixed parameters, sample sizes and percentages of zeros, various simulations are performed to assess the behavior of the estimators. Quantile residuals are defined to evaluate the assumptions of the proposed regression. Its usefulness is illustrated by an experiment conducted to assess the soil microbiology in a sugarcane field.

Effects of different legume green manures on tropical soil microbiology after corn harvest

Legume green manure is a popular practice for sustainable agriculture in tropical environments. The use of grain, forage and purely green manure legumes in a subhumid tropical area in Northeast Brazil was investigated to determine the total C and N contents and estimate C-microbial biomass, soil basal respiration, microbial and metabolic quotients, before and after green manures and after the following corn. It was hypothesized that, although all legumes would increase microbial biomass and activity, there would be different effects in function of the main economic usage of the legume. The experimental design evaluated the soil before and after harvest of seven legumes: (i) purely green manures – sunn hemp (Crotalaria juncea L.) and velvet bean (Stizolobium aterrimum Piper & Tracy); (ii) forage – Campo Grande stylo (a physical mixture of 80% of Stylosanthes captata Vog. and 20% of Stylosanthes macrocephala) and calopo (Calopogonium mucunoides Dev.) and (iii) grain – common bean (Phaseolus vulgaris L.), cowpea (Vigna unguiculata L.) and peanut (Arachis hypogaea L.), followed by corn crop. After corn harvest, soils had the same total C and N contents before legume seeding, but C-microbial biomass was significantly higher in soils with grain and purely green manures, which produced higher shoot and root dry mass. Soil basal respiration decreased after corn harvest for all legumes comparing to control treatments, but metabolic stress increased in soils with forage species. Despite this, microbial quotient indicated an efficiency of the microorganisms in degrading the residual organic matter of all green manures used in corn.

Количественные показатели микроорганизмов аллювиальных почв естественных биотопов южного склона Большого Кавказа

This article discusses the results of studies on the microbiology of alluvial soils on the southern slope of the Greater Caucasus. The first information on the number of microorganisms in alluvial soils of natural biotopes is presented. The changes are associated with alluvial (river) sediments of the Shinchay River; the riverbed and above-floodplain terrace are compared. The analysis of soils of meadow-herbaceous (gramineous) and undersized shrubs (wild rose, blackberry, tamarix) vegetation is presented. Differences were revealed in quantitative indicators and the percentage ratio of individual groups of microbiota biotopes. In conclusion, it is concluded that the alluvial deposits of Shinchai in the 0–30 cm soil layer contain 3119.23 thousand/g microorganisms, and in soil samples taken from the above-floodplain terrace in a similar 0–30 cm layer — 4406.53 thousand/g.

Remediation by means of EDTA of an agricultural calcareous soil polluted with Pb.

The dispersion of mine tailings affects ecosystems due to their high content of potentially toxic elements. Environmental risk increases when the soil impacted by tailings is used for agriculture; this use may result in health impacts. This study analyzes the feasibility of remediating a calcareous soil (used for maize cultivation) polluted with lead in the semiarid zone of Zimapán, México, by using EDTA as an extractant. Total geoavailable and bioaccessible concentrations in the gastric and intestinal phases were determined to evaluate lead availability and health risk. The soil was then washed with EDTA, and the geochemical fractionation (interchangeable, carbonates, Fe/Mn oxy-hydroxides, organic matter-sulfides, and residual) and impact on the mesophile bacteria and fungi/yeast populations were analyzed. The results showed total Pb concentrations up to 647 ± 3.50mg/kg, a 46% bioaccessible fraction (297 ± 9.90mg/kg) in the gastric phase and a 12.2% (80 ± 5mg/kg) bioaccessible fraction in the intestinal phase, indicating a health and environmental risk. Meanwhile, the geochemical fractionation before washing showed a Pb fraction mainly consisting of Fe/Mn oxy-hydroxides (69.6%); this reducible fraction may progressively increase its bioaccessibility. Geochemical fractionation performed in the washed soil showed differences from that determined before the treatment; however, the iron and manganese fraction, at 42.4%, accounted for most of the Pb. The soil microbiology was also modified by EDTA, with an increase in aerobic bacteria and a decrease in fungi/yeast populations. Although 44% total lead removal was achieved, corresponding to a final concentration of 363.50 ± 43.50mg/kg (below national and USEPA standards), washing with EDTA increased the soluble and interchangeable lead concentrations. Statistical analysis indicated a significant effect (p < 0.05) of EDTA on the soil's geochemical fractionation of lead.

Artificially intelligent soil quality and health indices for ‘next generation’ food production systems.

Currently, the lack of a universal soil quality index (SQI) limits adoption of such an approach and may hinder improvements to crop productivity and environmental sustainability. Some SQIs rely only on physicochemical characteristics, which are slow to change and thus have low sensitivity in predicting soil degradation in an appropriate timescale. In contrast, microorganisms respond quickly to changes in land-use and/or management. Furthermore, microbes generate the enzymes and biophysical structures required for many soil functions which thus drive ‘fertility’, ‘health’, and ‘quality’. Therefore, understanding of community-driven transformations should enable prediction of the trajectories of soil quality in response to management. However, the multitude of varied consequences and feedback loops which emerge dependent on site-specific factors are beyond the capability of models that currently exist. Enormous amounts of soil (meta)genomic data has been generated in the last decade. In parallel, advances in Artificial Intelligence (AI) have revolutionized our capacity to create predictive models in several areas, such as helping plant breeders searching for specific beneficial traits, and informing crop-management by predicting changes in the weather. As soil microbiologists and bioinformaticians, we contend that creating a universal, robust and dynamic Artificially Intelligent Soil Quality Index (AISQI) implies taking advantage of machine learning algorithms and soil microbiome data together with conventional physicochemical parameters and productivity data. This index must be flexible enough to encompass regional peculiarities but allow for comparative studies. Refining different models within the same index might improve its accuracy helping make real-time predictions. The establishment of a collaborative effort is fundamental to creating this index with maximum utility in enhancing agricultural management practices and ecosystem sustainability.

Biochar applications combined with paddy-upland rotation cropping systems benefit the safe use of PAH-contaminated soils: From risk assessment to microbial ecology

This study aimed to establish a method allowing the safe use of polycyclic aromatic hydrocarbon (PAH)-contaminated soils through the combination of biochar applications and different cropping systems. The impact of biochar applications under different cropping systems on the human health risks of PAHs and soil microbiology was elucidated. The residual PAHs were the lowest in rhizosphere soils amended with 2% corn straw-derived biochar pyrolyzed at 300 °C (CB300) under the paddy-upland rotation cropping (PURC) system. Human health risks resulting from the ingestion of PAH-contaminated carrot roots / rice grains under the PURC system were significantly lower than those under continuous upland cropping systems. The greatest diversity, richness and network complexity of soil microbial communities occurred under the PURC system combined with the 2% CB300 treatment. Soil microbial functions associated with soil health and PAH biodegradation were enhanced under this strategy, while the pathogen group was inhibited. Primarily owing to its high sorption capacity, bamboo-derived biochar pyrolyzed at 700 °C realized in the reduction of PAHs, but weakly influenced shifts in soil microbial communities. Overall, the combination of PURC systems and low-temperature-pyrolyzed nutrient-rich biochar could efficiently reduce the human health risks of PAHs and improve soil microbial ecology in agricultural fields.

Fungi and Bacteria in the Dark-Humus Forest Soil

A comparative assessment of the pools of fungal and bacterial biomass in dark humus forest soil has been performed using luminescent microscopy and cascade filtration. Cascade filtration indicates that the bacterial biomass is compatible to the fungal biomass (31–54% for bacteria and 46–69% for fungi) in the upper horizons of investigated soil. However, in the lower horizons, the bacterial biomass predominates (up to 69% at the depth of 100 cm). Thus, the cascade filtration method has made it possible to refine data on the bacterial biomass in the soil and to show for the first time that the biomass of bacteria is compatible to the biomass of fungi in the upper soil horizons and exceeds fungal biomass in the lower horizons. This method provides for a more accurate assessment of both the number and the biomass of bacteria, which allows revising the opinion of many soil microbiologists on the significant prevalence of fungal biomass in soils.

The analysis of methods of investigation of negative impact on mesofauna and microbiology of soils of contaminated solutions for foam makers

Nowdays the problem of fires is becoming more widespread and global in scale. According to statistics published by the Ukrainian Research Institute of Civil Protection obtained as a result of analysis of the fire registration cards of the SES of Ukraine – for 11 months of 2019 the number of fires in Ukraine increased by 23.1% compared to the same period last year. The most common extinguishing agent being used by fire and rescue units today is а foam. The main component of different foaming agents are surfactants. Their excessive emission into the environment leads to pollution of the surrounding areas. The constant increasing of the land areas affected by the effects of firefighting influence both the state of the environment in the region as a whole and the level of soil fertility. Microbiological indicators and the content of harmful substances in crops are increasing and thus the negative impact on public health grows. The analysis of microbiological monitoring methods with the aim of the contaminated solutions foam makers impact on the nearby territories’ soils estimation has been carried out. The pros and cons of modern physical and chemical methods of soils quality estimation have been defined. The necessity of microbiological methods usage which allow to get full and sufficient information not only about the pollution volume, but also to rate the results of such pollution has been justified. It is determined that in order to obtain the most complete and objective information about the microbiological condition of soils contaminated with foaming agents for fire extinguishing, a complex of bioindication methods should be used. It should include a growth test and a chromosome aberration test, with the use of test organisms that are most sensitive to the chemical composition of the test solution for firefighting.

Active projects for teaching and learning soil microbiology and applications of inoculants to increase perceived subject matter understanding and acquisition of knowledge.

Active methodologies for teaching propose tools and strategies for improving student learning by using participative and integrative approaches. These lead students to autonomous research for industry problems and solutions. This study aimed to apply active-project active methodologies to undergraduate soil microbiology and inoculant courses to verify students' perception of their knowledge levels on these topics. Forty undergraduate students received the traditional methodology that presented theoretical contents referring to the soil microbiology and inoculants; one group of twenty also elected to receive active methodologies based instruction during which they developed active projects that were structured in seven steps: briefing, bibliographic research, problematization and resolution, solutions, abstract and banner creation, and presentation. At the end of the academic year, all students answered a questionnaire to verify the perception of their levels of knowledge of soil microbiology and inoculants. Regarding the topic of microbial inoculants, perceived knowledge was the same for both groups, but overall, the active methodologies group had higher perceived knowledge of good practices of inoculation. The two groups were clustered by a multivariate approach, confirming that the use of active projects can increase the knowledge and level of subject matter understanding. The active projects contributed to undergraduate students' increased assimilation and perceived understanding of soil microbiology subject matter content and microbial inoculant issues. The active projects can be explored in other subdivisions of soil science, including agriculture and environmental studies.

The Result of Second Field Test of Cellulose Degrading Bacteria from Mangrove Soil Microbiology Handout

Learning media is one of the most important component in a learning process, both inside or outside a classroom. In order to provide proper learning for students based on the researcher’s prior research, at least, three different field tests are needed. This research is a quasi-experiment, which is provide result of the second field test of cellulose degrading bacteria from mangrove soil microbiology handout both in cognitive and psychomotor ability. The research aims are to describe effect of this microbiology handout on student’s cognitive achievement and gain data about inappropriate laboratory written instruction in this handout. Data gained by conducting learning activities using developed handout which were 2 times in class theory and three times of practical activity using instuctions written in the handout. In the end of class theory activity, students did a posttest to confirm their improvement of cognitive ability while their practical ability was observed using observation sheet along with their activities. Results showed that handout which being tested could improve student cognitive at average level (n gain 0,419). In contrast, several parts of laboratory written instruction need to be fixed to make better microbiology handout.