Suppression of Brugia malayi (sub-periodic) larval development in Aedes aegypti (Liverpool strain) fed on blood of animals immunized with microfilariae

Preliminary studies were carried out to investigate the role of filarial specific antibodies, raised in an animal model against the filarial parasite, Brugia malayi (sub-periodic), in blocking their early development in an experimental mosquito host, Aedes aegypti (Liverpool strain). In order to generate filarial specific antibodies, Mongolian gerbils, Meriones unguiculatus, were immunized either with live microfilariae (mf) of B. malayi or their homogenate. Mf were harvested from the peritoneal cavity of Mongolian gerbils with patent infection of B. malayi and fed to A. aegypti along with the blood from immunized animals. Development of the parasite in infected mosquitoes was monitored until they reached infective stage larvae (L3). Fewer number of parasites developed to first stage (L1) and subsequently to L2 and L3 in mosquitoes fed with blood of immunized animals, when compared to those fed with blood of control animals. The results thus indicated that filarial parasite specific antibodies present in the blood of the immunized animals resulted in the reduction of number of larvae of B. malayi developing in the mosquito host.

Evaluation of adenosine deaminase seric activity in the diagnosis of bovine tuberculosis

Determination of seric levels of adenosine deaminase (ADA), an enzyme produced by monocytes/macrophages and lymphocytes, has been used in the diagnosis of human tuberculosis (TB). In the present study, ADA seric activity was evaluated comparatively to the comparative tuberculin test in the diagnosis of bovine tuberculosis. Two hundred fifty-six cattle were classified by origin and by the comparative tuberculin test as TB-positive animals (n = 52, from herds where the Mycobacterium bovis had previously been isolated), and TB-negative animals (n = 204, TB-free herds). The mean ADA seric value from the TB-positive group (4.45 ± 2.33 U/L) was significantly lower (p = 0.008) than that observed in sera from the TB-negative group (6.12 ± 4.47 U/L). When animals from a herd with clinical cases of enzootic bovine leukosis of TB-negative group were withdrawn from analysis, the mean ADA seric values of TB-negative group (5.12 ± 3.75 U/L) was not significantly different anymore from that of the TB-positive group (p = 0.28). There was no agreement in the diagnosis of bovine TB between comparative tuberculin test and determination of ADA seric values, using two different cutoff points, being 6.12 U/L and 15.0 U/L, (kappa = -0.086 and kappa = -0.082, respectively). In conclusion, the determination of ADA seric activity was not a good auxiliary test for bovine TB, because it was not able to distinguish between TB-positive and TB-negative animals.

Bovine papillomavirus type 2 detection in the urinary bladder of cattle with chronic enzootic haematuria

The bovine papillomavirus type 2 (BPV-2) involvement in the aetiology of chronic enzootic haematuria associated to bracken fern ingestion has been suggested for a long time. However, a few reports have shown the presence of the BPV-2 in urinary bladder tumors of cattle. The aim of this study was to investigate the presence of the BPV-2 infection in the urinary bladder of cattle with chronic enzootic haematuria in Brazilian cattle herds. Sixty-two urinary bladders were collected from adult cattle in beef herds from the north region of the state of Paraná, Brazil. According to clinical and pathological finds the specimens were distributed in three groups: the group A was constituted by 22 urinary bladders with macroscopic lesions collected at necropsy of cattle with clinical signs of chronic enzootic haematuria; the group B by 30 urinary bladders with macroscopic lesions collected in a slaughterhouse of cows coming from bracken fern-endemic geographical region; and the group C (control) by 10 urinary bladders without macroscopic lesions collected from asymptomatic cattle in a bracken fern-free geographical region. By a semi-nested polymerase chain reaction (PCR) assay, with an internal control, a fragment of the BPV-2 L1 gene with 386 bp length was amplified in 36 (58%) urinary bladder. The rate of BPV-2 positive urinary bladders was 50% (11/22) for group A, 80% (24/30) for group B, and 10% (1/10) for group C (control). The rate of the positive results found in groups A and B that included urinary bladder samples with macroscopic lesions was 67% (35/52) and the detection of the BPV-2 in both groups was significantly higher (P < 0.05) than in the control group. RFLP with Rsa I and Hae III enzymes evaluated the specificity of the BPV-2 amplicons. The PCR internal control that amplified a 626 bp fragment of the ND5 gene of the bovine mitochondrial genome was amplified in all analyzed samples and excluded false-negatives or invalid results in the semi-nested PCR. These results suggest the BPV-2 involvement in the chronic enzootic haematuria aetiology and open the perspective of the development of new strategies for the control of this disease that is the major cause of economical losses in beef herds from many Brazilian geographical regions.

Biology of three species of the Meccus phyllosomus complex (Hemiptera: Reduviidae: Triatominae) fed on blood of hens and rabbits

Aspects related to hatching, life time, number of blood meals to molt, mortality, feeding time and postfeed defecation delay for each instar of Meccus phyllosomus, M. mazzottii, and M. bassolsae, life-cycle were evaluated and compared in two cohorts of each of those three species, fed on hens or rabbits. No significant (p > 0.05) differences were recorded among cohorts fed on hens respect to cohorts fed on rabbits in M. phyllosomus and M. mazzottii and the average time of hatching was 21.5 days for cohorts fed on hens and 22.5 for cohorts fed on rabbits. Average egg-to-adult development times were no significant (p > 0.05) different between both cohorts of M. phyllosomus and M. mazzotti, independent of the blood meal source. The average span in days for each instar fed on hens was not significantly different to the average span for each instar fed on rabbits, when comparisons were made by species. The number of blood meals at each nymphal instar varied from 1 to 6 in both cohorts of each species. The mortality rates were higher on older nymphs, in both cohorts of M. phyllosomus and M. bassolsae, whereas they were higher on first instar nymphs on M. mazzottii. Mean feeding time was no significant (p > 0.05) different in triatomines fed on hens or fed on rabbits, when each species were compared separately. A similar number of nymphs of each cohort, completed the cycle. Defecation delay was no significant (p > 0.05) different when cohorts fed on hens and fed on rabbits were compared by species. Most of the studied parameters showed no significant (p > 0.05) differences among those cohorts fed on hens and for fed on rabbits, which could mean a high degree of association of those species with birds as much as mammals, under wild conditions, increasing their capacity to colonize human dwellings.

Hepatic stereology of schistosomiasis mansoni infected-mice fed a high-fat diet

High-fat diets induce weight gain and fatty liver in wild-type mice. Schistosomiasis mansoni infection also promotes hepatic injury. This study was designed to quantify hepatic alterations in schistosomiasis mansoni-infected mice fed a high fat-rich chow compared to mice fed a standard rodent chow, using stereology. Female SW mice fed each either high-fat diet (29% lipids) or standard chow (12% lipids) over 8 months, and then were infected with Schistosoma mansoni cercariae. Four experimental groups were studied: infected mice fed a high-fat diet (IHFC) or standard chow (ISC), uninfected mice fed a high-fat diet (HFC) or standard chow (SC). Mice were sacrificed during early infection (9 weeks from exposure). The following hepatic biometry and the stereology parameters were determined: volume density (hepatocytes [h], sinusoids [s], steatosis [st] and hepatic fibrosis [hf]); numerical density (hepatocyte nuclei - Nv[h]); absolute number of total hepatocyte N[h], normal hepatocyte N[nh], and binucleated hepatocyte N[bh], percentage of normal hepatocyte P[nh] and binucleated hepatocyte P[bh]. IHFC and HFC groups exhibited TC, HDL-C, LDL-C, and body mass significantly greater (p < 0.05) than control group. No significant differences were found regards liver volume (p = 0.07). Significant differences were observed regards P[nh] (p = 0.0045), P[bh] (p = 0.0045), Nv[h] (p = 0.0006), N[h] (p = 0.0125), N[bh] (p = 0.0164) and N[nh] (p = 0.0078). IHFC mice group presented 29% of binucleated hepatocytes compared to HFC group (19%), ISC group (17%) and SC (6%). Volume density was significantly different between groups: Vv[h] (p = 0.0052), Vv[s] (p = 0.0025), Vv[st] (p = 0.0004), and Vv[hf] (p = 0.0007). In conclusion, schistosomiasis mansoni infection with concurrent high-fat diet promotes intensive quantitative changes in hepatic structure, contributing to an increasing on hepatic regeneration.

Isolation and identification of bovine tuberculosis in a Brazilian herd (São Paulo)

Mycobacterium was verified in animals from a Brazilian dairy herd, a total of 42 samples from 30 cows were submitted to culture and the isolated strains were analyzed by two polymerase chain reaction (PCR), the first specific for species belonging to the Mycobacterium complex (MTBC) and the other for differentiating M. tuberculosis from M. bovis. Twenty seven samples (64.3%) from 18 animals (60%) were positive for mycobacteria by culture, including samples from 15 retrofaryngeal lymphnodes (55.5%), 9 prescapular lymphnodes (33.3%), 2 lungs (7.4%), and 1 liver (3.7%). All isolated colonies were confirmed by PCR to contain MTBC organisms, and were identified as M. bovis by the same methodology.

Biology of three species of North American Triatominae (Hemiptera: Reduviidae: Triatominae) fed on rabbits

Aspects related to hatching, lifetime, number of blood meals for molting, mortality, feeding time, and postfeeding defecation delay were evaluated and compared in each instar of three North American Triatominae: Triatoma gerstaeckeri, Triatoma lecticularia and Triatoma protracta, all of them fed on rabbits. No significant differences (p > 0.05) were found among the three species regarding mean hatching rate, which was close to 20 days. Egg-to-adult development times were significantly shorter (p < 0.05) for T. lecticularia. Number of blood meals for molting to next instar ranged from one to five for T. protracta, and from one to six for T. gerstaeckeri and T. lecticularia. Mortality rates were higher in younger nymphs of T. lecticularia and T. protracta, while rates in T. gerstaeckeri were higher in fifth-instar nymphs. Mean feeding time was longest in T. gerstaeckeri, followed by T. lecticularia. More than twice the number of T. gerstaeckeri nymphs completed the development process, if compared to the nymphs from the other two species. Defecation delay was less than 10 min for T. lecticularia, T. protracta and the youngest nymphs of T. gerstaeckeri. Results point out that these three species may be important potential vectors of Trypanosoma cruzi for human populations, in areas of Mexico where these species are currently present.

Effect of untreated bed nets on blood-fed Phlebotomus argentipes in kala-azar endemic foci in Nepal and India

Observational studies in the Indian subcontinent have shown that untreated nets may be protective against visceral leishmaniasis (VL). In this study, we evaluated the effect of untreated nets on the blood feeding rates of Phlebotomus argentipes as well as the human blood index (HBI) in VL endemic villages in India and Nepal. The study had a "before and after intervention" design in 58 households in six clusters. The use of untreated nets reduced the blood feeding rate by 85% (95% CI 76.5-91.1%) and the HBI by 42.2% (95% CI 11.1-62.5%). These results provide circumstantial evidence that untreated nets may provide some degree of personal protection against sand fly bites.

Bovine tuberculosis in Doñana Biosphere Reserve: the role of wild ungulates as disease reservoirs in the last Iberian lynx strongholds.

Doñana National Park (DNP) in southern Spain is a UNESCO Biosphere Reserve where commercial hunting and wildlife artificial feeding do not take place and traditional cattle husbandry still exists. Herein, we hypothesized that Mycobacterium bovis infection prevalence in wild ungulates will depend on host ecology and that variation in prevalence will reflect variation in the interaction between hosts and environmental risk factors. Cattle bTB reactor rates increased in DNP despite compulsory testing and culling of infected animals. In this study, 124 European wild boar, 95 red deer, and 97 fallow deer were sampled from April 2006 to April 2007 and analyzed for M. bovis infection. Modelling and GIS were used to identify risk factors and intra and inter-species relationships. Infection with M. bovis was confirmed in 65 (52.4%) wild boar, 26 (27.4%) red deer and 18 (18.5%) fallow deer. In the absence of cattle, wild boar M. bovis prevalence reached 92.3% in the northern third of DNP. Wild boar showed more than twice prevalence than that in deer (p<0.001). Modelling revealed that M. bovis prevalence decreased from North to South in wild boar (p<0.001) and red deer (p<0.01), whereas no spatial pattern was evidenced for fallow deer. Infection risk in wild boar was dependent on wild boar M. bovis prevalence in the buffer area containing interacting individuals (p<0.01). The prevalence recorded in this study is among the highest reported in wildlife. Remarkably, this high prevalence occurs in the absence of wildlife artificial feeding, suggesting that a feeding ban alone would have a limited effect on wildlife M. bovis prevalence. In DNP, M. bovis transmission may occur predominantly at the intra-species level due to ecological, behavioural and epidemiological factors. The results of this study allow inferring conclusions on epidemiological bTB risk factors in Mediterranean habitats that are not managed for hunting purposes. Our results support the need to consider wildlife species for the control of bTB in cattle and strongly suggest that bTB may affect animal welfare and conservation.

Molecular characterization of G and P-types bovine rotavirus strains from Goiás, Brazil: high frequency of mixed P-type infections

In this study, 331 samples from calves less than one month old from a dairy herd in the district of Piracanjuba, state of Goiás, Brazil were tested for rotavirus. Thirty-three samples (9.9%) tested positive for rotavirus. Out of those, 31 were submitted to G and P characterization by reverse transcription followed by semi-nested polymerase chain reaction. Two samples were characterized as G6P[1], three as G10P[11] and five as G6P[11]. The majority of the samples (51.6%) displayed multiple P genotypes (P-genotype mixtures), including typical human genotypes P[4] and P[6M], suggesting the occurrence of co-infections and genetic reassortment. Also, the detection of human genotypes in bovine samples may be considered evidence of the zoonotic potential of rotaviruses. To our knowledge, this is the first report of such a high frequency of P genotype mixtures in bovine rotavirus samples. It also increases data on G and P rotavirus genotypes circulating in dairy herds in Brazil and can help in the development of more efficient immunization approaches, thereby controlling infection and reducing economical losses.

Production of TNF-α, nitric oxide and hydrogen peroxide by macrophages from mice with paracoccidioidomycosis that were fed a linseed oil-enriched diet

Omega-3 polyunsaturated fatty acids (n-3 PUFA) can modulate the immune system and their primary effect is on macrophage function. Paracoccidioidomycosis (PCM) is an endemic systemic mycosis in Latin America that is caused by the dimorphic fungus Paracoccidioides brasiliensis (Pb). Macrophages are the main defence against this pathogen and have microbicidal activity that is dependent on interferon-Γ and tumour necrosis factor (TNF)-α. These cytokines stimulate the synthesis of nitric oxide (NO) and hydrogen peroxide (H2O2), leading to the death of the fungus. To study the effect of n-3 PUFA on the host immune response during experimental PCM, macrophages that were obtained from animals infected with Pb18 and fed a diet enriched by linseed (LIN) oil were cultured and challenged with the fungus in vitro. The macrophage function was analysed based on the concentrations of TNF-α, NO and H2O2. LIN oil seems to influence the production of TNF-α during the development of disease. A diet enriched with LIN oil influences the microbicidal activity of the macrophages by inducing the production of cytokines and metabolites such as NO and H2O2, predominantly in the chronic phase of infection.

IL-10 release by bovine epithelial cells cultured with Trichomonas vaginalis and Tritrichomonas foetus

Trichomonas vaginalis and Tritrichomonas foetus are parasitic protists of the human and bovine urogenital tracts, respectively. Several studies have described the cytotoxic effects of trichomonads on urogenital tract epithelial cells. However, little is known about the host cell response against trichomonads. The aim of this study was to determine whether T. foetus and T. vaginalis stimulated the release of the cytokine interleukin (IL)-10 from cultured bovine epithelial cells. To characterise the inflammatory response induced by these parasites, primary cultures of bovine oviduct epithelial cells were exposed to either T. vaginalis or T. foetus. Within 12 h after parasite challenge, supernatants were collected and cytokine production was analysed. Large amounts of IL-10 were detected in the supernatants of cultures that had been stimulated with T. foetus. Interestingly, T. vaginalis induced only a small increase in the release of IL-10 upon exposure to the same bovine cells. Thus, the inflammatory response of the host cell is species-specific. Only T. foetus and not T. vaginalis induced the release of IL-10 by bovine oviduct epithelial cells.

Safety and immunomodulatory effects of three probiotic strains isolated from the feces of breast-fed infants in healthy adults: SETOPROB study.

We previously described the isolation and characterization of three probiotic strains from the feces of exclusively breast-fed newborn infants: Lactobacillus paracasei CNCM I-4034, Bifidobacterium breve CNCM I-4035 and Lactobacillus rhamnosus CNCM I-4036. These strains were shown to adhere to intestinal mucus in vitro, to be sensitive to antibiotics and to resist biliary salts and low pH. In the present study, a multicenter, randomized, double-blind, placebo-controlled trial with 100 healthy volunteers in three Spanish cities was carried out to evaluate the tolerance, safety, gut colonization and immunomodulatory effects of these three probiotics. Volunteers underwent a 15-day washout period, after which they were randomly divided into 5 groups that received daily a placebo, a capsule containing one of the 3 strains or a capsule containing a mixture of two strains for 30 days. The intervention was followed by another 15-day washout period. Patients did not consume fermented milk for the entire duration of the study. Gastrointestinal symptoms, defecation frequency and stool consistency were not altered by probiotic intake. No relevant changes in blood and serum, as well as no adverse events occurred during or after treatment. Probiotic administration slightly modified bacterial populations in the volunteers' feces. Intestinal persistence occurred in volunteers who received L. rhamnosus CNCM I-4036. Administration of B. breve CNCM I-4035 resulted in a significant increase in fecal secretory IgA content. IL-4 and IL-10 increased, whereas IL-12 decreased in the serum of volunteers treated with any of the three strains. These results demonstrate that the consumption of these three bacterial strains was safe and exerted varying degrees of immunomodulatory effects.

Effects of Crypthecodiniumcohnii, Chlorela spp. e Isochrysisgalbana addition to milk replacer on goat kids and lamb growth.

The effects of Crypthecodinium cohnii (Cryp.), Chlorela spp. (Chlo.) and Isochrysis galbana (Iso.) addition to milk replacer on goat kids and lambs growth were evaluated. About 80 Majorera goat kids (males and females) and 80 Canarian sheep lambs were randomly assigned into four different groups (by specie) according to diet. Control groups were fed with a commercial milk replacer at 16% (w/w); Cryp. groups received a commercial milk replacer (15.1% w/w) supplemented with 9 g of a paste of C. cohnii; Chlo. groups received a commercial milk replacer (15.1% w/w) supplemented with 9 g of a paste of Chlorela spp.; Iso. groups received a commercial milk replacer (15.1% w/w) supplemented with 9 g of a paste of I. galbana. After colostrum period, animals were individually bottle-fed twice daily (8 am and 8 pm) ad libitum with the corresponding diet until day 60 of life. Animals were weighted every week at 8 am and liquid diet intake was recorded weekly. No effects of microseaweed addition were observed, neither growth nor milk replacer intake.

Histological behavior of glutaraldehyde cross-linked bovine collagen injected into the human bladder for the treatment of vesicoureteral reflux

Endoscopic subureteral collagen injection has become an accepted means for the treatment of vesicoureteral reflux in children. The aim of this study was to evaluate the histological behavior of glutaraldehyde cross-linked bovine collagen implants. The specimens were harvested from 29 patients who underwent reimplant surgery 2 to 30 months (mean 9.5) after unsuccessful subureteral injection therapy. In addition to routine hematoxylin and eosin staining, a new staining method (solophenyl red 3BL) able to demonstrate selectively neoformation of types I and III human collagen, was applied. Invasion of host fibroblasts into the bovine implant and the formation of endogenous types I and III collagen were demonstrated in all 29 cases. Adverse histological reactions were rare and, if present, they were predominantly of an inflammatory nature.