Size Does Matter: Application-driven Approaches for Soil Metagenomics

Metagenomic analyses can provide extensive information on the structure, composition, and predicted gene functions of diverse environmental microbial assemblages. Each environment presents its own unique challenges to metagenomic investigation and requires a specifically designed approach to accommodate physicochemical and biotic factors unique to each environment that can pose technical hurdles and/or bias the metagenomic analyses. In particular, soils harbor an exceptional diversity of prokaryotes that are largely undescribed beyond the level of ribotype and are a potentially vast resource for natural product discovery. The successful application of a soil metagenomic approach depends on selecting the appropriate DNA extraction, purification, and if necessary, cloning methods for the intended downstream analyses. The most important technical considerations in a metagenomic study include obtaining a sufficient yield of high-purity DNA representing the targeted microorganisms within an environmental sample or enrichment and (if required) constructing a metagenomic library in a suitable vector and host. Size does matter in the context of the average insert size within a clone library or the sequence read length for a high-throughput sequencing approach. It is also imperative to select the appropriate metagenomic screening strategy to address the specific question(s) of interest, which should drive the selection of methods used in the earlier stages of a metagenomic project (e.g., DNA size, to clone or not to clone). Here, we present both the promising and problematic nature of soil metagenomics and discuss the factors that should be considered when selecting soil sampling, DNA extraction, purification, and cloning methods to implement based on the ultimate study objectives.     

Maximum slat width for cooling efficiency of horticultural produce in wooden crates

The influence of wood package design on airflow distribution was investigated for forced-air cooling using horticultural produce simulators. The position of grooves on the container walls was tested using slat width of 100–200 mm and airflow rates ranging from 0.0005 to 0.003 m³ kg⁻¹ s⁻¹. The package opening configurations were compared based on their impact on the energy added to the system using a methodology previously developed. For this purpose, apples and sweet corns were taken as examples of produce from two different extremes in the respiration activity range. For airflow rates as low as 0.0005 m³ kg⁻¹ s⁻¹one groove at the bottom of the container produced a cooling process more uniform than the other one-groove configurations and even two grooves because of natural convection effect. If packing low respiration rate produce, increasing airflow rate could compromise the process energy efficiency because of air circulation obstruction for less vented containers. For high respiration rate produce enlarging open area above 2.4% would be recommended rather than increasing airflow rate to enhance cooling energy efficiency.     

Differentiation of soil fauna in desert agriculture of the Mariut region

Summary The soil fauna of 23 sites in the Mariut region, west of Alexandria, Egypt, was sampled monthly or seasonally for periods of 1,5–2.5 years. These sites range from unutilized littoral sand dunes, to free-grazing pasture, to low- and high-intensity dry farming, to irrigated agriculture. The average population densities of the encountered taxa were compared by corresponding analysis and by ascending hierarchic classification. With intensification of the agricultural practices, true detritivores of the original system, capable of dealing with the available plant and animal litter and adapted to the harsh desert climate, are replaced by other mesic detritivores adapted to the changes in litter quality (less animal and more plant) and also adapted to the new physical and chemical soil conditions. Meanwhile, the increase in plant biomass encourages the appearance and proliferation of cryptic phytophages (agricultural pests), occupying the niche of the removed large (domestic) herbivores. Carnivores (e.g.Carabidae) also change accordingly. Sites can thus be characterized on the basis of their soil faunas, according to the stage of their evolution, and a system of exchange of missing key taxa between sites should be established.     

Seasonal differentiation of soil mesofauna in a littoral dune of the Egyptian Mediterranean coast

Summary Soil mesofauna (sensu Ghilarov, i.e., larger than 1 mm) was sampled from under four shrub species (Lygos raetam, Thymelaea hirsuta, Lycium shawii, and Josonia candicans) growing on the littoral oolitic sand dunes at Gharbaniat, 53 km W of Alexandria, Egypt, for a period of more than 30 months, spanning 3 years. Population density (PD) was calculated in relation to area of shrub canopy. Seasons were considered, according to the prevailing climatic conditions as well as to earlier physiological studies on the animals concerned, as: four winter months, four summer months and two months for each of the transitional seasons spring and autumn. The PD values obtained for each season thus defined were combined for the 41 taxa that were sampled. The table of these data was treated by correspondence analysis (CA) and ascending hierarchic classification (AHC), by the Roux DATA-VISION programme (suitable for Apple). Results from such treatment show that fauna as related to shrub species are differentiated along the first factorial axis, while as related to seasons they are differentiated along the second axis. Taxa common to all shrubs and to all seasons are centrally placed and cause some confusion in the resolution. Summer taxa are more distantly placed than those related to the other three seasons. Fauna under Lygos is richer, more diverse, and more complementary as regards trophic levels (with predators), and could probably be the source for replenishing the fauna of the other shrubs at all seasons. The AHC method did not show a similarly clear differentiation.Dedicated to the late Prof. Dr. M.S. Ghilarov     

Rates of decomposition of plant residues and available nitrogen in soil, related to residue composition through simulation of carbon and nitrogen turnover

The dynamics of inorganic N in soil following the application of plant residues depends on their composition. We assumed that all plant materials are composed of similar components, each decomposing at a specific rate, but differ in the proportions of the various components. The NCSOIL model that simulates C and N turnover in soil was used to link the rates of residue decomposition to their composition, defined as soluble, cellulose-like and lignin-like C and N, and thereby integrate short and long-term effects of residues on available N dynamics in soil. Five plant residues in a wide range of C:N ratios were incubated in soil for 24 weeks at 30 °C, during which C and N mineralization were measured. The materials with large C:N ratios (corn, rice hulls and wheat straw) were also incubated with NH₄⁺-N to avoid N deficiency. The residues were analyzed for total and soluble C and N. The partitioning of insoluble C and N between cellulose- and lignin-like pools was optimized by best fit of simulated C and N mineralization to measured results. The decomposition rate constants of the soluble and lignin-like pools were assumed to be 1.0 and 10⁻⁵ d⁻¹, respectively, and that of the cellulose-like pool, obtained by model optimization against mineralization of cellulose with NH₄⁺-N in soil, was 0.051 d⁻¹. The optimized, kinetically defined lignin-like pool of all residues was considerably larger than lignin contents normally found in plant residues by the Van Soest procedure. Gross N mineralization of tobacco and rape residues was similar, but N recovery from tobacco was larger, because a larger fraction of its C was in the lignin-like pool. N in rice hulls, corn and wheat residues was mostly recalcitrant, yet rice hulls did not cause N deficiency, because most of its C was recalcitrant too. The soluble components of the residues had strong short-term effects on available N in soil, but the cellulose-like pool was equally important for short and medium-term effects. Soluble and cellulose-like C were 29 and 42% of total C, respectively, in corn and 7 and 50% in wheat. Maximal net inorganic N losses, measured in both residue treatments after 2 weeks, were 42 mg g⁻¹ C applied as corn and 31 mg g⁻¹ C applied as wheat, or 84 and 110 mg g⁻¹ decomposed C of corn and wheat, respectively. Rice hulls immobilized N slowly, but by the end of 24 weeks all three residues immobilized 26-27 mg N kg⁻¹ C applied. The different dynamics of N immobilization demonstrated the need to determine the decomposability of C and N rather than their total contents in plant residues.     

Factors Affecting Fertility According to the Postpartum Period in Crossbred Dual-Purpose Suckling Cows in the Tropics

To determine the relationship between first service conception rate (FSC) and the postpartum period (PP), 6160 calvings from 2250 crossbred dual-purpose cows were analysed. The effects of breed predominance (BT: Bos taurus and BI: Bos indicus), season of service (dry, intermediate and humid), parity (1 or 2 and more), agroecological region (Sub-humid and Dry Tropical Forest) and milk production were studied. The postpartum period was grouped according to the calving–first service interval as G1 ≤ 60 days (n = 1889); G2 > 60 ≤ 90 days (n = 1436); G3 > 90 ≤ 120 days (n = 1030); and G4 > 120 days (n = 1805). Data were analysed by chi-squared test. The overall mean for FSC was 58.2% and was lower in G1 (p < 0.01). All PP showed a breed effect on FSC, with mean values of 61.6% and 53.3% for BI and BT, respectively. Season of service also affected FSC, which was significantly lower during the humid period: 51.2% versus 57.4% and 61.5% for the intermediate and dry periods (p < 0.01), respectively. Parity and environmental effects were not observed on FSC. Levels of milk yield did influence the FSC, which decreased from 66.7% and 61.0% for milk yields of < 1500 kg and between 1501 and 2000 kg to 46.0% in milk yields > 2000 kg (p < 0.01).     

Favus in a fighting cock caused by Microsporum gallinae

The first diagnosis of favus in chickens in Costa Rica was made in a 1-year-old fighting cock that had lesions surrounding the comb. The etiologic agent was isolated and identified as Microsporum gallinae. The rooster recovered during topical treatment with tolnaftate and oral treatment with griseofulvin.     

Epidemiological aspects of filariosis in dogs on the coast of Paraná state, Brazil: with emphasis on Dirofilaria immitis

The present study determined the prevalence and geographical distribution of Dirofilaria immitis and other filariae, from dogs in littoral areas of Paraná state, in Brazil. This survey spanned eight months, between 1998 and 1999, and was also designed to compare the efficacy of different tests for diagnosis of heartworm infection in that area. Blood samples were collected from 256 native-owned dogs distributed along the Paraná coastal area. Five diagnostic procedures were used: direct smear examination, the Knott's modified test, filtration assay, and two heartworm antigen detection kits. A follow-up imaging exam was performed to support the heartworm diagnosis. The imaging diagnosis included radiographic and ultrasonographic exams of six dogs that had positive results for the heartworm antigen detection kits, but showed different microfilarial burdens. The presence and severity of radiographic and ultrasonographic signs were compared with the results obtained in microfilariae detection and antigen tests. Diagnostic parasitology results indicated that 31.25% of the dogs were microfilaremic. Three different microfilariae were recovered: D. immitis, Dipetalonema reconditum, and the third (mf3) was not identified. D. reconditum was the species with the highest prevalence: 22.6%. In general, D. immitis prevalence was 5.47% (28.57% occult infections), but it varied along the coast and the range was from 0 to 20%. No correlation could be established between the overall scores for microfilarial counts (small or large numbers) and the severity of radiographic results or the likelihood of detecting filariae in the pulmonary artery using echocardiography. The finding of a different type of microfilaria (mf) suggested the existence of a third species in Paraná state, whose prevalence was 4.68%. These results show that to obtain a reliable diagnosis of heartworm infection, antigen detection kits are indicated. Knott's test or filtration should be performed to confirm microfilaremia and not for diagnosis of heartworm infection. Imaging tests support parasitology exams and add more about severity of infection. The northern areas, specially Guaraque?aba and Ilha das Pe?as, presented the highest number of heartworm-infected dogs.     

An approach to identify dairy cows being responsive to recombinant bovine somatotropin

Forty-eight primiparous and pregnant Holstein cows at 202 (+/-59) days in milk and average daily milk production (DMP) of 22 (+/-5.3) kg at the beginning of the trial were used to study the possibility of identifying animals that are more responsive to recombinant bovine somatotropin (rbST). Animals were housed in free-stall barns, milked three times a day and fed a total mixed ration. Animals were injected with one dose (500 mg each) of rbST every 21 days during a 105-day total period (five doses in all). Daily milk production was recorded to measure average apparent response (AAR) and peak of apparent response (PAR). Average apparent response was defined as the average increase in milk production, expressed as kilogram per cow per day, for days 0-18 after dosing, as compared to the average production calculated for days -1, -2, -3, 19, 20 and 21 after dosing. Peak of apparent response was calculated as the increase in milk production for each day from 0 to 18 days after dosing as compared to the average production for days -1, -2 and -3. There was a positive linear correlation (r = 0.608; p = 0.0001) between AAR for the first 21 days and AAR for the total 105-day period (AATR). A correlation coefficient was calculated between each individual PAR value and AATR. The highest correlation coefficient (r = 0.511; p = 0.0002) was observed for days 8, 9 and 10, and therefore these days were considered the best estimates for calculating AAR when DMP records are not available. Also, high somatic cells count had a negative impact on milk yield response to treatment (p = 0.0162).     

Global biodiversity conservation priorities

The location of and threats to biodiversity are distributed unevenly, so prioritization is essential to minimize biodiversity loss. To address this need, biodiversity conservation organizations have proposed nine templates of global priorities over the past decade. Here, we review the concepts, methods, results, impacts, and challenges of these prioritizations of conservation practice within the theoretical irreplaceability/vulnerability framework of systematic conservation planning. Most of the templates prioritize highly irreplaceable regions; some are reactive (prioritizing high vulnerability), and others are proactive (prioritizing low vulnerability). We hope this synthesis improves understanding of these prioritization approaches and that it results in more efficient allocation of geographically flexible conservation funding.