Earthworm influence on carbon dioxide and nitrous oxide fluxes from an unfertilized corn agroecosystem

Earthworms modify the soil environment through their feeding, casting, and burrowing activities, which may lead to more decomposition and respiration in aerobic microsites and more denitrification in anaerobic microsites. The objective of this study was to determine whether earthworms increase CO₂ and N₂O fluxes from an unfertilized corn agroecosystem. Earthworm populations within field enclosures (2.9 m²) were reduced by repeatedly applying carbaryl insecticide, then single and mixed populations of Lumbricus terrestris L. and Aporrectodea caliginosa (Savigny) were added. Gas samples were collected once a week for 14 weeks, from June to September 2005. Carbaryl applications reduced, but did not eliminate earthworms from enclosures. The CO₂ and N₂O fluxes were affected by the sampling date, with peak gas fluxes after rainfall events. Mean CO₂ and N₂O fluxes during the study period tended to be greater from enclosures with added earthworms than the control (no earthworms added), but were not significantly affected by earthworm treatments due to the low survival rate of introduced earthworms. Better control of earthworm populations in the field is required to fully assess the impact of earthworms on CO₂ and N₂O fluxes from temperate agroecosystems.     

Bacterial load reduction of live prey for fish larval feeding using Ox-Aquaculture©

Reduction of the bacterial load in live prey is paramount to achieving efficient microbiological control during fish larval rearing. Several methods have been tested using physical and chemical disinfection procedures. Nevertheless, chemicals are difficult to deal with because of the frequent problems encountered when disposing of their residual products and because of the sensitivity of the live prey. Hydrogen peroxide is a good disinfectant that has been used for a long time in other fields of research and decomposes in non‐toxic products. However, it has not been applied for disinfection in aquaculture until recent years. The effects of a hydrogen peroxide‐based product, Ox‐Aquaculture^©, on rotifers and Artemia nauplii survival and on their associated microbial population have been tested in the present study, a disinfection protocol is proposed, and its effects on live prey fatty acids composition after enrichment were checked. More than 80% survival and a 90% reduction of total heterotrophic bacteria and Vibrionaceae were obtained in rotifers exposed for 15 min to 40 mg L^?1 of the product. In the case of Artemia nauplii, 90% survival and 94.5% reduction of heterotrophic bacteria were obtained after 5 min exposure to 8000 mg L^?1 with a further 82.8% reduction of Vibrionaceae. No differences in the fatty acid composition of the live prey were detected after disinfection with hydrogen peroxide. No oxidation of polyunsaturated fatty acids was observed. A short time exposure and easy removal of the product from the treated live prey make hydrogen peroxide an interesting chemical for industrial application.     

Tolerance to Formalin by a Fluidized-Bed Biofilter and Rainbow Trout Oncorhynchus mykiss in a Recirculating Culture System

Formalin tolerances of a fluidized-bed sand biofilter and rainbow trout Oncorhynchus mykiss were tested in a semi-closed freshwater recirculating culture system. Progressively higher formalin levels were tested until fish mortality (±1.9%) occurred. A 1-h exposure to 167 ppm formalin, followed by flushing, was safe for trout (no mortality) at 15.0 C but not at 16.5 C. A 110-ppm indefinite treatment (no flushing) was safe at 17.3 C, but 120 ppm at 17.3 C and 100 ppm at 17.8 C were not. Biofilter nitrification was not impaired by 1-h formdin treatments up to 167 ppm at 16.5 C, and not, usually, by indefinite treatments up to 120 ppm at 17.3 C. However, a find indefinite treatment of 70 ppm was followed by abnormally high nitrite levels for 9 d. Formaldehyde remained detectable in the system for 11 h during indefinite treatment at 120 ppm formalin. Tests on the system's Co₂ stripper did not indicate that it removed formaldehyde.     

Spatio‐temporal distribution of sea turtle strandings and factors contributing to their mortality in south‐eastern Brazil

1. Data on stranded sea turtles were examined between 2010 and 2016 along the northern region of Rio de Janeiro state and between 2016 and 2017 in the southern region, looking for spatio‐temporal patterns and determining which factors contributed to their mortality.
2. A total of 12,162 strandings of all five species that occur in Brazil were recorded, with Chelonia mydas being the most common (89.9%). Sea turtles use the Rio de Janeiro coast as a feeding and/or migration area. The intense upwelling (October to April) may be an important factor for the sea turtles feeding in this region, mainly for Eretmochelys imbricata and Dermochelys coriacea, which had a higher number of strandings during this period.
3. Areas further north of the study area include an important nesting site for Caretta caretta in Brazil, which explains the higher concentration of strandings of subadults/adults of this species in this region and during its nesting season.
4. Many anthropogenic threats to sea turtles were documented, mainly incidental capture in fisheries and marine debris, indicating possible hotspots for these threats in the regions of Sepetiba and Guanabara Bays, Cabo Frio, and São Francisco de Itabapoana.
5. Among the natural causes of strandings, the primary factors were chronic illness, endoparasites, and fibropapillomatosis. However, pollution may also be an indirect threat, which negatively affects these animals through reduced health and immunosuppression, leaving them more susceptible to opportunistic diseases.
6. These data are valuable for directing and implementing specific and local mitigation measures along the Rio de Janeiro state coast, such as avoiding bycatch hotspots through fleet communication programmes and/or area and seasonal closures, enforceable legislation, effective penalties and proper waste management.

     

Population declines of a long-lived salamander: a 20+-year study of hellbenders, Cryptobranchus alleganiensis

Accurate assessment of whether long-lived species are stable or declining is challenging. Life history characteristics such as delayed maturity result in relatively slow population responses to perturbations, so data should be collected across a relatively long time span. Because differential effects on age classes can be important, studies should also examine potential changes in the population's age structure. Moreover, multiple populations should be studied to indicate whether changes are regional or are restricted to local populations. We incorporated all three factors (long duration, multiple populations, age structure data) into our study of the conservation status of a long-lived aquatic salamander, the hellbender, Cryptobranchus alleganiensis. Over the 20+ years of this study, populations of hellbenders declined by an average of about 77%. This decline was characterized by a shift in size (age) structure, with a disproportionate decrease in numbers of young individuals. The change in density and age structure was consistent for populations in five rivers and for two subspecies (C. a. alleganiensis and C. a. bishopi), indicating that the decline is not restricted to one or two local populations. For the population with the most extensive data, the decline had clearly begun by the 1980s and there was a significant decrease in body condition over the period of the study. It is not known whether population declines for hellbenders have a single cause or whether each population has experienced independent declines.     

Development of a multiplex assay for the detection of antibodies to Borrelia burgdorferi in horses and its validation using Bayesian and conventional statistical methods

Lyme disease is a zoonotic, vector-borne disease and occurs in mammals including horses. The disease is induced by infection with spirochetes of the Borrelia burgdorferi sensu lato group. Infection of mammalian hosts requires transmission of spirochetes by infected ticks during tick bites. Lyme disease diagnosis is based on clinical signs, possible exposure to infected ticks, and antibody testing which is traditionally performed by ELISA and Western blotting (WB). This report describes the development and validation of a new fluorescent bead-based multiplex assay for the detection of antibodies to B. burgdorferi outer surface protein A (OspA), OspC and OspF antigens in horse serum. Testing of 562 equine sera was performed blindly and in parallel by using WB and the new multiplex assay. Because a true gold standard is missing for Lyme antibody testing, we performed and compared different statistical approaches to validate the new Lyme multiplex assay. One approach was to use WB results as a 'relative gold standard' in ROC-curve and likelihood-ratio analyses of the new test. Cut-off values and interpretation ranges of the multiplex assay were established by the analysis. The second statistical approach used a Bayesian model for the calculation of diagnostic sensitivities and specificities of the multiplex assay. The Bayesian analysis takes into consideration that no true gold standard exists for detecting antibodies to B. burgdorferi and estimated sensitivities and specificities of both tests that were compared. Therefore, the Bayesian analysis also resulted in an evaluation of diagnostic sensitivity and specificity of WB. Overall, the new assay was characterized by low background values and a wide dynamic quantification range for the detection of antibodies to OspA, OspC and OspF antigens of B. burgdorferi. The diagnostic sensitivity and specificity for the OspA bead-based assay were calculated as 49% and 85%, respectively, and by a standard ROC curve analysis only because the Bayesian model could not be run on this parameter. The Bayesian-derived diagnostic sensitivities of the OspC and OspF assays were 80% and 86%, respectively. For comparison, the Bayesian-derived estimates for WB resulted in sensitivities of 72% for OspC and 80% for OspF. The Bayesian diagnostic specificities of the multiplex assay were 79% and 69% for OspC and OspF, respectively. WB analysis had specificities of 92% for OspC and 77% for OspF. Although the analysis of a new assay in the absence of a true gold standard remains challenging, the approach used here can help to address this problem when new technologies and traditionally used test standards differ significantly in their analytical sensitivities, which consequently causes problems in the calculation of diagnostic sensitivity and sensitivity values for the new assay. In summary, the new multiplex assay for the detection of antibodies to B. burgdorferi OspA, OspC and OspF antigens in horse serum has improved analytical and diagnostic sensitivities compared to WB analysis. Multiplex analysis is a valuable quantitative tool that simultaneously detects antibodies indicative for natural infection with and/or vaccination against the Lyme pathogen.     

Sequential Elution Technique in Moss Pseudoscleropodium purum: Comparison between the Commonly Used Extracellular Extractant NiCl2 and Other New Extractants

The sequential elution technique (SET) is used to determine the distribution of elements in the different cell fractions of mosses. The extracellular extractants most commonly used in this technique are NiCl₂ and EDTA, although there are certain disadvantages associated with their use. In order to avoid such problems, we searched for new extractants that displace extracellularly bound metals, either because they are present at high concentrations (Ca) or because they have a high affinity for cation exchange sites (Hg and Au). The compounds HCl, NaCl, and CaCl₂ were tested as extractants of the former type, as possible alternatives for the determination of extracellular metals in the moss Pseudoscleropodium purum. Calcium chloride was finally chosen as is it potentially the most successful in terms of binding to the cation exchange sites without altering the membrane permeability. The concentration chosen was 160 mM, as this yielded maximum displacement of Zn without membrane alterations. An experiment was then carried out to test the efficiency of Ca in extracting extracellular Zn, under laboratory and field conditions. In addition, Hg and Au were tested, at different concentrations, as extractants with high affinity for cation exchange sites, as neither of these elements has previously been used in the SET, and both display electronic characteristics that suggest their potential usefulness in displacing other cations from cation exchange sites. The results obtained show that extraction of extracellularly bound metal by high concentrations of Ca should be ruled out, as total extraction of Zn was not achieved. Both Hg and Au produced membrane alterations at low concentrations and moreover, neither was more efficient at extracting Zn than the reference extractant (20 mM NiCl₂).     

Sex chromosomes behavior and G‐banding treatment of male meiosis in nuptial gift‐giving spiders of the family Trechaleidae

Preliminary genetic studies in Trechaleidae spider family show high variation in sex chromosomes and high heterocigocity, suggesting high chromatin plasticity. The trechaleids Paratrechalea ornata, Trechalea bucculenta and Trechaleoides biocellata are present in Uruguay. Males offering nuptial gifts during courtship have been reported in P. ornata and T. bucculenta but not in T. biocellata. Nuptial gifts are an inherited trait probably highly affected by environmental factors, which play an important role in gene expression. We hypothesize that this trait could be associated with tissue‐specific genes existing in G‐bands. We investigate the male meiosis in these 3 species, their sex chromosome system and the effects of G‐banding on their chromosomes, and elucidate genetic differences among them. Meiotic stages of the 3 species were submitted to Giemsa‐staining and G‐banding treatments. We observed a haploid number of n= 11 in P. ornata and n= 13 in both T. bucculenta and T. biocellata. Males from the 3 species presented an X₁X₂0 sex chromosome system, which is suggested as ancestral in Araneae. In P. ornata and T. bucculenta, both sex chromosomes were together and aligned in parallel until the segregation during anaphase I. In contrast to these species, sex chromosomes of T. biocellata usually remained distant from each other until diakinesis when they were observed associated in parallel disposition. Interstitial G‐bands were similar in P. ornata and T. bucculenta, and they both differed from those in T. biocellata. The special behavior of sex chromosomes in T. biocellata as well as the different G‐banding pattern of this species suggests the existence of novel modifications in this species.     

Accuracy assessment and screening of a dairy herd with paratuberculosis by three different ELISAs

Although the culture of Mycobacterium avium subsp. paratuberculosisis is the gold standard for the diagnosis of paratuberculosis, this bacterium is difficult to grow. In contrast, serological tests like ELISAs are inexpensive, rapid, and easy to perform. The aims of this study were to evaluate the accuracy of three different ELISAs: one with the commercial antigen PPA-3, another one with L5P (a recently described lipopentapeptide), and a third one with an in-house antigen whole cell lysates (WCL) of M. avium (MAA) strain D4ER (Study 1), and to compare them with other tests for paratuberculosis (PTB) diagnosis (Study 2). In Study 1, the sensitivities of the three ELISAs tested were 74.1%, 37% and 74.1%, respectively, whereas their specificities were 98.9%, 100% and 100%, respectively. In Study 2, we compared the three above-mentioned ELISAs with the intradermal reaction test using Avian PPD (PPDa) and fecal culture associated with Ziehl-Neelsen stain and PCR tests, in a dairy herd with 4.6% of cows with clinical signs of PTB. The results showed that fecal samples from 14 cows (16%) were culture-positive and that fecal samples from nine cows (10%) were PPDa-positive. Most of these animals (culture-positive and PPDa-positive) were detected as positive with any of the three ELISAs tested. Serological results showed that 31% of the animals were positive to ELISA-PPA-3, 17% to ELISA-L5P and 42.5% to ELISA-WCL. The combination of these three ELISAs identified 50.6% of the animals as positive in the infected herd. In particular, the results show that the locally developed ELISA seems to be useful for identifying many infected animals in a herd.