Biometrical, biological, biochemical and molecular characteristics of Meloidogyne incognita isolates and related species

Meloidogyne incognita is one of the most polyphagous species of root-knot nematodes occurring in Brazil and worldwide. Eight M. incognita isolates were studied, representing two enzymatic phenotypes (esterase and malate desydrogenase: I1/N1, I2/N1) and four cryptic Meloidogyne sp.1 (S2/N1) isolates, representing one cytological type (3n?=?40–46). Three M. hispanica isolates (Hi3/N1, 2n?=?32–36) and two of an atypical Meloidogyne sp.2 (S2a/N3, 3n?=?40–44) were included in this study for comparison. All isolates were tested with three M. incognita-specific molecular markers. The primer pairs B06F/R, miF/R and incK14F/R amplified three species-specific fragments of 1,200?bp, 955?bp and 399?bp, respectively for M. incognita and Meloidogyne sp.1 isolates. No amplification occurred in the M. hispanica and Meloidogyne sp.2 isolates, except with primers miF/R (1,650?bp). The genetic variability of the Meloidogyne spp. isolates was evaluated, using RAPD and ISSR markers. The phylogenetic analyses revealed two strongly supported monophyletic clades: clade I, consisting of M. hispanica and the atypical Meloidogyne sp.2 isolates, and clade II, clustering together all M. incognita and the Meloidogyne sp.1 isolates. Considering the biometrical, cytological and molecular approaches, it was possible to conclude that the isolates with three enzymatic phenotypes (I1/N1, I2/N1 and S2/N1) presented the characteristics described for M. incognita. Some correlations were detected between the isozymatic phenotypes and the tree topology (S2a/N3, Hi3/N1, I1/N1, S2/N1), but no strict correlation could be observed for the phenotype I2/N1 and one isolate of S2/N1. Morphologically, the Msp.2 isolates differ from M. incognita and M. hispanica by the female stylet features presenting straight cone tip and round pear shaped knobs, posteriorly sloping. The results of this study suggested that the Msp.2 isolates with phenotypes S2aN3 belong to a new or an unidentified species closely related to M. hispanica.     

Mapping genomic regions for reproductive traits in beef cattle: Inclusion of the X chromosome

Although the second largest chromosome of the genome, the X chromosome is usually excluded from genome-wide association studies (GWAS). Considering the presence and importance of genes on this chromosome that are involved in reproduction, the aim of this study was to evaluate the effect of its inclusion in GWAS on reproductive traits (scrotal circumference [SC], early pregnancy [P16] and age at first calving [AFC]) in a Nelore herd. Genotype data from 3,263 animals with the above-mentioned phenotypes were used. The results showed an increase in the variances explained by the autosomal markers for all traits when the X chromosome was not included. For SC, there was an increase of more than 10% for the windows on chromosomes 2 and 6. For P16, the effect was increased by almost 20% for windows on chromosome 5. The same pattern was found for AFC, with an increase of more than 10% for the most important windows. The results indicate that the noninclusion of the X chromosome can overestimate the effects of autosomes on SC, P16 and AFC not only because of the additive effect of the X chromosome itself but also because of its epistatic effect on autosomal genes.     

Description of outcomes of experimental infection with feline haemoplasmas: Copy numbers, haematology, Coombs’ testing and blood glucose concentrations

The aim of this study was to compare blood copy, haematological and glucose values between cats experimentally infected with either Mycoplasma haemofelis (Group HF: 10 cats), ‘Candidatus M. haemominutum’ (Group HM: 3 cats) or ‘Candidatus M. turicensis’ (Group TU: 3 cats). Blood samples were collected regularly up to 85 days post-infection (DPI) for haemoplasma real-time quantitative PCR, haematology, Coombs’ testing and blood glucose measurement. Statistical analysis was performed using a general linear model (ANOVA) appropriate for a repeated measures experiment with significance set as P < 0.05. Cats in Group TU had significantly lower blood copy numbers than cats in Group HF (P < 0.001) and HM (P < 0.001). All Group HF cats developed anaemia (often severe), macrocytosis and evidence of erythrocyte-bound antibodies whereas Groups HM and TU cats did not. Group HF had significantly lower PCVs, haemoglobin concentrations and red blood cell counts, and significantly higher mean cell volumes, than Groups HM and TU. In Group HF, erythrocyte-bound antibodies reactive at 4 °C (both IgM and IgG) appeared between 8 and 22 DPI and persisted for two to four weeks, whereas those reactive at 37 °C (primarily IgG) appeared between 22 and 29 DPI and persisted for one to five weeks. In most cats antibodies appeared after the fall in haemoglobin started. Although Group TU had significantly lower glucose concentrations than Groups HF (P = 0.006) and HM (P = 0.027), mean blood glucose concentrations remained within the reference range in all groups. This study demonstrates that M. haemofelis infection, in contrast to ‘Candidatus M. haemominutum’ and ‘Candidatus M. turicensis’ infection, can result in a severe macrocytic anaemia and the development of cold and warm reactive erythrocyte-bound antibodies.     

Gross and histologic evidence of sharp and blunt trauma in North Atlantic right whales (Eubalaena glacialis) killed by vessels.

Vessel-whale collision events represented the ultimate cause of death for 21 (52.5%) of the 40 North Atlantic right whales (Eubalaena glacialis) necropsied between 1970 and December 2006. Injuries seen in vessel-struck whales fall into two distinct categories: 1) sharp trauma, often resulting from contact with the propeller, and 2) blunt trauma, presumably resulting from contact with a vessel's hull. This study analyzes four trauma cases that resulted from vessel-whale collisions, which together provide a framework for a more critical understanding of lethal blunt and sharp trauma resulting from vessel collisions with right whales. In case no. 1, contact with a propeller resulted in three deep lacerations. The animal survived acute trauma only to succumb nearly 14 years later when the lesions reopened and became infected. In case no. 2, anecdotal reports linked the laceration of large arteries of the peduncle and histologic evidence of perimortem trauma at a bone fracture site to vessel-whale collision trauma. Case no. 3 had a laceration of the oral rete and a fracture of the rostrum. Both of the areas displayed histologic evidence of perimortem blunt trauma. Finally, in case no. 4, an antemortem mandibular fracture, two additional skull fractures, and widespread hemorrhage were consistent with severe blunt trauma. Evidence from each case, including the timing of trauma relative to the time of death and identifying characteristics of both trauma types, are presented. Before this study, no detailed comparative analysis of trauma pathology that resulted from lethal interactions between vessels and right whales had been conducted. This study demonstrates the importance of detailed gross and histologic examination in determining the significance and timing of traumatic events. This work represents a new paradigm for the differential diagnosis of lethal sharp and blunt trauma in right whales hit by ships and will enhance the present understanding of the impact of anthropogenic mortality on this critically endangered species.     

Acaricidal properties of the essential oil and precocene II obtained from Calea serrata (Asteraceae) on the cattle tick Rhipicephalus (Boophilus) microplus (Acari: Ixodidae)

Calea serrata Less. (Asteraceae), an endemic species of south Brazil known as "quebra-tudo", is used in Afro-Brazilian religious rituals and in folk medicine for treating liver disorders. Phytochemical studies of the n-hexane extract of this plant demonstrated the presence of precocene II, a benzopyran derivative known for its insecticidal activity. The aim of this work was to isolate this benzopyran and determine the chemical composition of the essential oil of C. serrata and further to evaluate the acaricidal activity of the essential oil and precocene II against the larvae of Rhipicephalus (Boophilus) microplus. The LC(99.9) and LC(50) values obtained with the oil, which presents precocene II and sesquiterpenes, were 3.94 μL/mL and 0.28 μL/mL, respectively. For precocene II this values were 4.25mg/mL and 1.78 mg/mL, respectively. The results indicate a synergistic interaction between the components of the oil and precocene II.     

Across-breed validation study confirms and identifies new loci associated with sexual precocity in Brahman and Nellore cattle

The aim of this study was to identify candidate regions associated with sexual precocity in Bos indicus. Nellore and Brahman were set as validation and discovery populations, respectively. SNP selected in Brahman to validate in Nellore were from gene regions affecting reproductive traits (G1) and significant SNP (p ≤ 10^–3) from a meta-analysis (G2). In the validation population, early pregnancy (EP) and scrotal circumference (SC) were evaluated. To perform GWAS in validation population, we used regression and Bayes C. SNP with p ≤ 10^–3 in regression and Bayes factor ≥3 in Bayes C were deemed significant. Significant SNP (for EP or SC) or SNP in their ±250 Kb vicinity region, which were in at least one discovery set (G1 or G2), were considered validated. SNP identified in both G1 and G2 were considered candidate. For EP, 145 SNP were validated in G1 and 41 in G2, and for SC, these numbers were 14 and 2. For EP, 21 candidate SNP were detected (G1 and G2). For SC, no candidate SNP were identified. Validated SNP and their vicinity region were located close to quantitative trait loci or genes related to reproductive traits and were enriched in gene ontology terms related to reproductive success. These are therefore strong candidate regions for sexual precocity in Nellore and Brahman.     

Multiplex real-time PCR SYBR Green for detection and typing of group III Clostridium botulinum

Clostridium botulinum type C and type D belonging to the group III organisms, are mainly responsible for animal botulism outbreaks. Clinical signs alone are often insufficient to make a diagnosis of botulism and a laboratory confirmation is required. Laboratory confirmation can be performed by demonstrating the presence of botulinum neurotoxins in serum, gastrointestinal contents, liver, wound of sick or dead animals, or by demonstrating the presence of C. botulinum in gastrointestinal contents, liver, and wound. Demonstration of spores in gastrointestinal contents or tissue of animals with clinical signs indicative of botulism reinforces the clinical diagnosis. With the aim of detecting and typing C. botulinum group III organisms, a multiplex real-time PCR SYBR Green was developed and in-house validated. Selectivity, limit of detection, relative accuracy, relative specificity, relative sensitivity, and repeatability of the method were investigated. The multiplex real-time PCR SYBR green used showed a 100% selectivity, 100% relative accuracy, 100% relative specificity, 100% relative sensitivity and a limit of detection of 277 and 580 DNA copies for C. botulinum type C and C. botulinum type D, respectively. The method reported here represents a suitable tool for laboratory diagnosis of type C and D botulism and for testing a large number of samples collected during the animal botulism surveillance and prevention activities.