Genetic and antigenic characterization of the matrix protein of two genetically distinct ovine lentiviruses

Small Ruminant Lentiviruses (SRLV) are a group of non-oncogenic retroviruses including Maedi-Visna virus (MVV) and Caprine Arthritis-Encephalitis virus (CAEV), which cause a chronic, multisystemic disease in sheep and goats, respectively. Phylogenetic analyses of SRLV are based in most cases on partial pol sequences. Several reports indicate that the species specificity of these viruses is not as strict as previously thought; MVV-like viruses have been found in goat populations and vice versa. Recently, the sequencing of some Italian ovine isolates has shown the presence of a new cluster more similar to classical caprine isolates (CAEV-like). Few data are available on the variability of structural proteins involved in the antibody response of infected animals. In this study, the gag gene of two genetically distinct ovine isolates, namely the MVV-like It-561 and the CAEV-like It-Pi1, was sequenced and the epitopes of matrix protein (MA) were mapped. Recombinant MAs and their subunits from both ovine aforementioned strains were tested against a panel of sheep and goat sera. Reactive epitopes were found in all three subunits of MA, although the central subunit displayed a more consistent reactivity. Epitope mapping of this subunit demonstrated that the amino acid sequence of at least one immunodominant epitope was quite different in the two strains. This antigenic variability may affect the sensitivity of a single strain-based immunoassay and suggests that both SRLV genotypes should be used in the development of future diagnostic tests, to avoid viral strain selection during the eradication programmes.     

Genetic and antigenic characterization of the surface lipoprotein P48 of Mycoplasma bovis

The presence of a membrane lipoprotein homologous to the P48 of Mycoplasma agalactiae was investigated in different Mycoplasma bovis isolates selected by geographical locations and biological properties. Its potential as a diagnostic tool was also discussed. The presence of a specific signal observed in all M. bovis field isolates probed with a rabbit antiserum raised against the M. agalactiae recombinant P48 demonstrated that this protein is structurally and antigenically conserved within the M. bovis cluster. No signal was detected when testing six different mycoplasma species found in cattle. The p48 gene was identified by PCR approach and partially sequenced. Full length gene sequence was obtained by direct bacterial chromosome sequencing. Five UGAs were selectively mutated into UGG and the full length mutated gene, lacking the signal peptide, was cloned and expressed in Escherichia coli. The purified recombinant antigen (r-P48) was evaluated as a potential marker of infection using a panel of 86 well-characterized sera from experimentally and naturally infected cattle. Specific IgM antibodies were detected within 6-9 days after experimental infection followed by an IgG response lasting from the third/fourth week after contact. Although antibody titers were well below those observed in sheep or goats infected with M. agalactiae, results suggest that M. bovis r-P48 can be used as a specific marker of infection.     

Detection of bacterial wilt infection caused by Ralstonia solanacearum in potato (Solanum tuberosum L.) through multifractal analysis applied to remotely sensed data

Potato bacterial wilt, caused by the bacterium Ralstonia solanacearum race 3 biovar 2 (R3bv2), affects potato production in several regions in the world. The disease becomes visually detectable when extensive damage to the crop has already occurred. Two greenhouse experiments were conducted to test the capability of a remote sensing diagnostic method supported by multispectral and multifractal analyses of the light reflectance signal, to detect physiological and morphological changes in plants caused by the infection. The analysis was carried out using the Wavelet Transform Modulus Maxima (WTMM) combined with the Multifractal (MF) analysis to assess the variability of high-resolution temporal and spatial signals and the conservative properties of the processes across temporal and spatial scales. The multispectral signal, enhanced by multifractal analysis, detected both symptomatic and latently infected plants, matching the results of ELISA laboratory assessment in 100 and 82%, respectively. Although the multispectral method provided no earlier detection than the visual assessment on symptomatic plants, the former was able to detect asymptomatic latent infection, showing a great potential as a monitoring tool for the control of bacterial wilt in potato crops. Applied to precision agriculture, this capability of the remote sensing diagnostic methodology would provide a more efficient control of the disease through an early and full spatial assessment of the health status of the crop and the prevention of spreading the disease.     

Small ruminant lentiviruses in Jordan: evaluation of sheep and goat serological response using recombinant and peptide antigens

Small ruminant lentiviruses infect sheep and goats worldwide, causing chronic progressive diseases and relevant economic losses. Disease eradication and prevention is mostly based on serological testing. The goal of this research was to investigate the presence of the small ruminant lentiviruses (SRLVs) in Jordan and to characterize the serological response in sheep and goat populations. A panel of sera were collected from flocks located in Northern Jordan and Jordan Valley. The samples were tested using three ELISA assays: a commercially available ELISA based on p25 recombinant protein and transmembrane peptide derived from British maedi–visna virus (MVV) EV1 strain, an ELISA based on P16-P25 recombinant protein derived from two Italian strains representative of MVV- and caprine arthritis encephalitis virus (CAEV)-like SRLVs, and an ELISA based on SU5 peptide from the same two Italian isolates. The results indicate that both MVV- and CAEV-like strains are present in Jordan and that the majority of the viruses circulating among sheep and goat populations belong to the MVV-like genotype.     

Leukocyte response and phagocytic activity in Nile tilapia experimentally infected with <Emphasis Type="Italic">Enterococcus</Emphasis> sp.

This study evaluated the total and differential leukocyte counting and the phagocytic activity in Nile tilapia Oreochromis niloticus experimentally injected with Enterococcus sp. in the swim bladder. Fish were distributed in four treatments in triplicates of non-injected fish, fish injected with 1 ml of sterile saline solution 0.65%, and fish injected with 1 × 10³ and 1 × 10⁶ colony-forming units (CFU) of Enterococcus diluted in 1 ml sterile saline. Twenty-four hours after injection, the fish were anesthetized and the blood collected for white blood cell (WBC) counts, differential counting of WBC, and phagocytic activity of blood leukocytes. The increased numbers of WBC and lymphocytes were followed by decreased number of monocyte after infection. The percentages of phagocytic activities in the blood were 55.3 and 55.9%, respectively, in tilapia injected with 1 × 10³ and 1 × 10⁶ CFU/ml.     

Autochthonous probiotic as growth promoter and immunomodulator for Astyanax bimaculatus cultured in water recirculation system

This work aimed to evaluate the probiotic Lactobacillus spp. in the microbiota of Astyanax bimaculatus reared in a water recirculation system. The experiment was randomized, using 800 postlarvae distributed into eight polyethylene containers, separated into two treatments, in quadruplicate: supplementation with Lactobacillus spp. (Probiotic) and no supplementation (Control). After 90 days of culture, 13 fish per container were anaesthetized. Three of these underwent microbiological evaluation, and haematological analyses were carried out on five. Immunological assessment was performed on another five fish. Zootechnical parameters were evaluated for all animals. In the microbiological evaluation, the probiotic group presented higher counts of lactic acid bacteria, but lower counts of Vibrio spp., Pseudomonas spp., and Staphylococcus spp., when compared with control. After haematological analysis, the supplemented group presented low haematimetric indices of Mean Corpuscular Haemoglobin Concentration and Mean Corpuscular Haemoglobin (MCHC and MCH, respectively) and higher amounts of total leucocytes, thrombocytes and circulating monocytes when compared with control. The immunological profile did not differ between treatments. In the zootechnical performance, probiotic group presented higher productivity, survival and apparent food efficiency when compared with control. Therefore, when Lactobacillus spp. was supplemented in the diet of A. bimaculatus, immunocompetence and the zootechnical performance of animals increased.     

Serological characterization of the new genotype E of small ruminant lentivirus in roccaverano goat flocks

Maedi visna virus (MVV) and caprine arthritis encephalitis virus (CAEV) are a heterogeneous group of infectious agents affecting sheep and goats. Due to their natural cross-species infection they are referred to as small-ruminant lentiviruses (SRLV). Recently a new genetic cluster, highly divergent from MVV and CAEV was identified in the north-west part of Italy. A panel of genotype E specific antigens was developed and evaluated in flocks infected with B1 and E strains. The results clearly indicate that a strain specific antigen is required to correctly identify animals infected with different genotypes.     

Genetic immunization with the immunodominant antigen P48 of Mycoplasma agalactiae stimulates a mixed adaptive immune response in BALBc mice

A DNA vaccine against contagious agalactia was developed for the first time, encoding the P48 of Mycoplasma agalactiae. Specific immune responses elicited in BALB/c mice were evaluated. Both total IgG and IgG1 were detected in mice vaccinated with pVAX1/P48. Proliferation of mononuclear cells of the spleen, levels of gamma interferon, interleukin-12, and interleukin-2 mRNAs were enhanced in immunized animals. Results indicate that pVAX1/P48 vaccination induced both T_h1 and T_h2 immune responses. Nucleic acid immunization could be a new strategy against M. agalactiae infections and may be potentially used to develop vaccines for other Mycoplasma diseases.