Early development of Betta splendens under stereomicroscopy and scanning electron microscopy

Betta splendens is a very important ornamental species. The current paper describes the embryonic and larval development of B. splendens under stereomicroscopy and scanning electron microscopy. Eggs and larvae from natural spawning were collected at different developmental stages at previously established intervals and analysed. The eggs of B. splendens are yellowish, clear, spherical, demersal, translucent and telolecithal with a large amount of yolk. Between 0-2 h post-initial collection (hpIC), the eggs were at the egg cell, first cleavage and morula stages. The blastula stage was identified at 2-3 hpIC and the early gastrula phase was observed at 3-4 hpIC with 20% epiboly, which was finalized after 13-18 hpIC. When the pre-larvae were ready to hatch, the appearance of somites and the free tail were observed, at 23-25 hpIC. At 29 hpIC, the majority of larvae had already hatched at an average temperature of 28.4 +/- 0.2 degrees C. The newly hatched larvae measured 2.47 +/- 0.044 mm total length. The mouth opened at 23 h post-hatching (hPH) and the yolk sac was totally absorbed at 73 hPH. After 156 hPH, the heart was pumping blood throughout the entire larval body. The caudal fin, operculum and eyes were well developed at 264 hPH. When metamorphosis was complete at 768 hPH, the larvae became juveniles. The current study presents the first results about early development of B. splendens and provides relevant information for its reproduction, rearing and biology.

Induced reproduction and early development histology of Oscar Astronotus ocellatus (Agassiz, 1831)

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Caracterização das proteínas caveolinas -1 e -2 na placenta de conceptos bovinos clonados transgênicos

The transgenic application of green fluorescent protein (GFP) as fetal cell marker on cattle cloned placenta could provide an exclusive model for studying the morphologic and immunologic maternal-fetal interactions, providing information about its mapping, distinguishing the fetal from maternal cells. This model will have direct application, mainly because these animals present problems during its development. With this model's support, we intend to verify the substances transport between mother and fetus during endocytosis, through the immunolocalization of protein named caveolae. For these, we used 06 cloned bovine and 30 cattle samples of artificial insemination (AI) with 90 days of pregnancy, which had been their development interrupted by humanitarian slaughter of the recipient and recovery of the pregnant uterus. We collected the placentome and the chorion. A part of the samples was cut and fixed, by immersion, on a solution containing 4% of parafomaldehyde or 10% of formaldehyde on a sodium phosphate buffer (PBS), at 0,1 M pH 7.4, Zamboni solution (4% of paraformaldehyde, 15% of picric acid, on sodium phosphate buffer 0,1 M pH 7.4), metacarn (60% of metanol, 30% of chloroform, and 10% glacial acetic acid), for morphologic and immunohistochemistry verification for caveolinas proteins -1 and -2 (CAV -1 and CAV-2). The caveolins -1 were found in fetal and maternal villi, but its strongest staining was observed in the endometrial stroma. The caveolins -2 had positive staining in trophoblast and chorioallantoic membrane, and specifically in giant trophoblastic binucleated cell. Therefore the results were compared between cloned cattle and from AI or natural mating, for assisting on detection of the reason of many placental alterations, embryonic losses, spontaneous abortion, post-natal mortality and large offspring syndrome on laboratory-manipulated animals. The result suggests that the proteins caveolins -1 and -2 (CAV-1 and CAV-2) are part of the caveolae composition and important structures related to the molecule transfer to the fetus, nourish it through endocytosis and pinocytosis.

Plasma Steroid Dynamics in Late- and Near-term Naturally and Artificially Conceived Bovine Pregnancies as Elucidated by Multihormone High-resolution LC-MS/MS

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Immunohistochemical approach to the pathogenesis of clinical cases of bovine Herpesvirus type 5 infections

Meningoencephalitis by Herpesvirus type 5 (BoHV-5) in cattle has some features that are similar to those of herpetic encephalitis in humans and other animal species. Human Herpesvirus 3 (commonly known as Varicella-zoster virus 1), herpes simplex viruses (HSV), and equid Herpesvirus 1 (EHV-1) induce an intense inflammatory, vascular and cellular response. In spite of the many reports describing the histological lesions associated with natural and experimental infections, the immunopathological mechanisms for the development of neurological disorder have not been established. A total of twenty calf brains were selected from the Veterinary School, University of São Paulo State, Araçatuba, Brazil, after confirmation of BoHV-5 infection by virus isolation as well as by a molecular approach. The first part of the study characterized the microscopic lesions associated with the brain areas in the central nervous system (CNS) that tested positive in a viral US9 gene hybridization assay. The frontal cortex (Fc), parietal cortex (Pc), thalamus (T) and mesencephalon (M) were studied. Secondly, distinct pathogenesis mechanisms that take place in acute cases were investigated by an immunohistochemistry assay. This study found the frontal cortex to be the main region where intense oxidative stress phenomena (AOP-1) and synaptic protein expression (SNAP-25) were closely related to inflammatory cuffs, satellitosis and gliosis, which represent the most frequently observed neurological lesions. Moreover, MMP-9 expression was shown to be localized in the leptomeninges, in the parenchyma and around mononuclear infiltrates (p < 0.0001). These data open a new perspective in understanding the role of the AOP-1, MMP-9 and SNAP-25 proteins in mediating BoHV-5 pathogenesis and the strategies of host-virus interaction in order to invade the CNS.

Influence of sex on basal and dickkopf-1 regulated gene expression in the bovine morula

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Genomic data as the hitchhiker's guide to cattle adaptation: tracking the milestones of past selection in the bovine genome

The bovine species have witnessed and played a major role in the drastic socio-economical changes that shaped our culture over the last 10,000 years. During this journey, cattle hitchhiked on human development and colonized the world, facing strong selective pressures such as dramatic environmental changes and disease challenge. Consequently, hundreds of specialized cattle breeds emerged and spread around the globe, making up a rich spectrum of genomic resources. Their DNA still carry the scars left from adapting to this wide range of conditions, and we are now empowered with data and analytical tools to track the milestones of past selection in their genomes. In this review paper, we provide a summary of the reconstructed demographic events that shaped cattle diversity, offer a critical synthesis of popular methodologies applied to the search for signatures of selection (SS) in genomic data, and give examples of recent SS studies in cattle. Then, we outline the potential and challenges of the application of SS analysis in cattle, and discuss the future directions in this field.

Transgenic cotton-fed Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae) affects the parasitoid encarsia desantisi Viggiani (Hymenoptera: Aphelinidae) development

Cotton cultivars expressing Cry proteins are widely used to control lepidopteran pests. The effects of transgenic plants containing insecticidal Cry proteins on non-target species must be comprehended for a better and rational use of this technology for pest management. We investigated the influence of the Bt cotton cultivars NuOPAL and FM 975 on biological parameters of Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae), a non-target pest of Bt cotton cultivars and on its parasitoid Encarsia desantisi Viggiani (Hymenoptera: Aphelinidae). The experiments were conducted in a climatized room, and the non-transgenic near isolines were used for rearing whiteflies as control hosts. The effects of the Bt cotton cultivars on the period of embryonic and larval development and the percentage of adult emergence of B. tabaci were assessed. The period required for embryonic, larval, and pupal development and the percentage of emergence and longevity of E. desantisi females were determined using Bt cotton-fed and non-Bt cotton-fed B. tabaci as hosts. Both Bt cotton cultivars resulted in a decrease of approximately 20% of adult emergence of B. tabaci. Differently, an increase of approximately 10% of adult emergence of E. desantisi was observed for parasitoids that used hosts fed with both Bt cotton cultivars. However, female parasitoid longevity decreased when their hosts were fed on Bt cotton cultivars. Our data suggest that the use of Bt cotton cultivars in association with the biological control agent E. desantisi could be functional for the management of B. tabaci in Bt cotton crops.

Comparison of Brucella agar, CITA and Farrel media for selective isolation of Brucella abortus from semen of bovine bulls

Brucellosis remains as a public health concern worldwide. In domestic animals, the disease is characterized by reproductive disorders in male and female. Besides extensive use of serological tests and recent development of molecular biology techniques, microbiological culture of Brucella species is yet considered a “gold standard” method for diagnosis. Here, semen of 335 bovine bulls was subjected simultaneously to microbiological culture in Brucella agar, Farrell media, and CITA media to evaluate comparatively the best selective media for isolation of Brucella sp. Among all 335 samples, B. abortus B19 strain was isolated from semen of five (1.49%) bulls using the three selective media. However, Farrell media was considered the best selective media for microbiological diagnosis, because of allowed isolation of B. abortus B19 strain from bull semen without bacterial commensal or fungal contamination of plates.

In vitro production of bovine embryos using Sigma antioxidant supplement®, α-tocopherol and L-ascorbic acid

This study tested the effect of Sigma antioxidant supplement®, α-tocopherol (vitamin E) and L-ascorbic acid (vitamin C) in the culture medium of bovine embryos. In experiment 1, in vitro produced bovine zygotes were cultured in Human Tubal Fluid (HTF): Eagle’s Basic Medium (BME) with: Group 1 – 50 µm vitamin C; Group 2 – 200 µm vitamin E; Group 3 – 25 µm vitamin C and 100 µm vitamin E; Group 4 – 1 µl/ml Sigma antioxidant supplement®; and the Control group – HTF:BME only. In experiment 2, embryos were cultured in high or low oxygen tension with HTF:BME + Sigma antioxidant supplement® or in HTF:BME alone (Control). The data were analyzed using ANOVA followed by Tukey’s test. The results of experiment 1 showed a negative effect (P < 0.05) of vitamin E on blastocyst production in Group 2 (19.7 ± 0.1%). This effect was reduced in Group 3 by the addition of vitamin C (26.1 ± 0.2%). The use of vitamin C alone (34.9 ± 0.3%) or the Sigma antioxidant supplement® (33.3 ± 0.7%) did not increase (P > 0.05) the number of blastocysts produced compared with the control group (30.1 ± 0.5%). During experiment 2, there was no effect (P > 0.05) from the culture medium or the O2 concentrations used, indicating that the reduction of the O2 concentration did not improve blastocyst production.

Development of a Model of Natural Infection with Mycobacterium bovis in White-Tailed Deer

The objective of this study was to develop a suitable experimental model of natural Mycobacterium bovis infection in white-tailed deer (Odocoileus virginianus), describe the distribution and character of tuberculous lesions, and to examine possible routes of disease transmission. In October 1997, 10 mature female white-tailed deer were inoculated by intratonsilar instillation of 2 3 103 (low dose) or 2 3 105 (high dose) colony forming units (CFU) of M. bovis. In January 1998, deer were euthanatized, examined, and tissues were collected 84 to 87 days post inoculation. Possible routes of disease transmission were evaluated by culture of nasal, oral, tonsilar, and rectal swabs at various times during the study. Gross and microscopic lesions consistent with tuberculosis were most commonly seen in medial retropharyngeal lymph nodes and lung in both dosage groups. Other tissues containing tuberculous lesions included tonsil, trachea, liver, and kidney as well as lateral retropharyngeal, mandibular, parotid, tracheobronchial, mediastinal, hepatic, mesenteric, superficial cervical, and iliac lymph nodes. Mycobacterium bovis was isolated from tonsilar swabs from 8 of 9 deer from both dosage groups at least once 14 to 87 days after inoculation. Mycobacterium bovis was isolated from oral swabs 63 and 80 days after inoculation from one of three deer in the low dose group and none of four deer in the high dose group. Similarly, M. bovis was isolated from nasal swabs 80 and 85 days after inoculation in one of three deer from the low dose group and 63 and 80 days after inoculation from two of four deer in the high dose group. Intratonsilar inoculation with M. bovis results in lesions similar to those seen in naturally infected white-tailed deer; therefore, it represents a suitable model of natural infection. These results also indicate that M. bovis persists in tonsilar crypts for prolonged periods and can be shed in saliva and nasal secretions. These infected fluids represent a likely route of disease transmission to other animals or humans.

Lesion Development in White-tailed Deer (Odocoileus virginianus) Experimentally Infected with Mycobacterium bovis

The recent discovery of tuberculosis in free-living white-tailed deer in northeastern Michigan underscores the need for increased understanding of the pathogenesis of tuberculosis in wildlife species. To investigate lesion development in white-tailed deer, 32 deer were experimentally infected by intratonsilar instillation of 300 colony-forming units of Mycobacterium bovis. Three deer each were euthanatized and examined at days 15, 28, 42, and 56 after inoculation, and five deer each were euthanatized and examined at days 89, 180, 262, and 328 after inoculation. Microscopic lesions first were seen in the medial retropharyngeal lymph node and lung 28 and 42 days after inoculation, respectively. Lung lesions were present in 12 (38%) of 32 deer, involving 23 lung lobes. Left caudal and right middle and caudal lobes were involved in 17 (74%) of the 23 affected lung lobes. Lesions in the medial retropharyngeal lymph node first appeared as granulomas composed of aggregates of macrophages and Langhans-type giant cells. Some early granulomas contained centrally located neutrophils. As granulomas developed, neutrophils were replaced with a central zone of caseous necrosis that first showed signs of mineralization 42 days after inoculation. Granulomas increased in size as the zone of caseous necrosis expanded. Peripheral fibrosis, first seen at 56 days after inoculation, progressed to only a thin fibrous capsule by 328 days after inoculation. By the termination of the study, the central necrotic core of the granuloma contained abundant liquefied necrotic material and grossly resembled an abscess. Although tuberculous lesions in white-tailed deer follow a developmental pattern similar to that in cattle, fibrosis is less pronounced and the advanced lesions may liquefy, a change seldom reported in cattle. An understanding of lesion development will aid in the identification of the spectrum of disease that may be seen in this important wildlife reservoir of tuberculosis.

A New Approach for Managing Bovine Tuberculosis: Veterinary Services’ Proposed Action Plan

Bovine tuberculosis (TB) is a serious disease with animal health, public health, and international trade consequences. The cooperative Federal-State-industry effort to eradicate bovine TB from cattle in the United States has made significant progress since the program’s inception in 1917. However, the goal of eradication remains elusive. This proposed action plan presents Veterinary Services’ (VS’) current thinking about changes we are considering for the TB program to address our current challenges. This action plan will: 1. Reduce the introduction of TB into the U.S. national herd from imported animals and wildlife by: o Applying additional requirements to cattle imports from Mexico o Enhancing efforts to mitigate risks from wildlife 2. Enhance TB surveillance by: o Crafting a comprehensive national surveillance plan o Accelerating diagnostic test development to support surveillance 3. Increase options for managing TB-affected herds by: o Conducting epidemiological investigations and assessing individual herd risk o Applying whole-herd depopulation judiciously and developing alternative control strategies o Applying animal identification (ID) standards to meet animal ID needs 4. Modernize the regulatory framework to allow VS to focus resources where the disease exists 5. Transition the TB program from a State classification system to a science-based zoning approach to address disease risk To succeed, this new approach will require VS’ continued partnership with State animal health and wildlife officials, other Federal agencies, industry, international partners, academia, and other stakeholders. Successful partnerships will allow us to use available resources efficiently to achieve program objectives and protect our nation’s herd. Implementation of the VS proposed action plan will benefit Federal and State animal health officials, the regulated industries, and producers by allowing a more rapid response that employs up-to-date science and can adapt rapidly to changing situations.

On the Ecology and Distribution of Echinococcus spp. (Cestoda: Taeniidae), and Characteristics of Their Development in the Intermediate Host. II. Comparative Studies on the Development of Larval E. multilocularis Leuckart, 1863, in the Intermediate Host

This paper reports the results of a comparative study of the development of the larval Echinococcus multilocularis Leuckart, 1863), and associated tissue reaction in naturally and experimentally infected mammals representing 31 species. The histogenesis of the larval cestode was traced in detail in arvicoline rodents of several species, and interspecific differences were defined. In arvicoline rodents, the developing larva exhibited host-specific characteristics within about a month after infection was established. The tissue reaction in Microtus oeconomus was characterized by the production of a large quantity of detritus around the larva, and by the formation of a thick epithelioid zone. In one subspecies, M. oeconomus innuitus, development of the larva was retarded, and the detrital mass was often calcified; in another, M. oeconomus operarius, the detritus rarely became calcified and the larva proliferated more rapidly. In M. pennsylvanicus, the tissue reaction was minimal, and little detritus was present. The characteristics of the tissue reaction in M. montebelli placed it in an intermediate position between the aforementioned species. In Clethrionomys rutilus, a thin epithelioid zone and an outer zone of loose collagenous fibers composed the adventitial layer; exogenous budding was retarded in this vole. A minimal tissue reaction occurred in Lagurus curtatus. In Lemmus spp., larger cysts were characteristic, but areas of small-cystic proliferation were always present. Similar differences in species or subspecies of Citellus and Dicrostonyx were described. Lesions of alveolar bydatid disease in man also were studied. The invasive growth of the larval cestode in the human liver involves a process comparable to small-cystic proliferation in the natural intermediate hosts. Although the later stages of development of the larval cestode are inhibited in man, exogenous proliferation of vesicles continues for the life of the host. The lesion in man was compared with a morphologically similar formation produced by anomalous development of the larval E. granulosus in the bovine liver. The latter is distinguished by the absence of areas of small-cystic proliferation. Non-echinococcal lesions found in the tissues studied, some of which resembled foci caused by the larval E. multilocularis, were briefly discussed.

Taking an Endemically Infected Bovine Tb Area Through to Livestock Clearance. A Case Study of West Taupo, New Zealand

The Pest Management Strategy for Bovine Tuberculosis (Tb) in New Zealand aims to achieve efficient freedom from Tb by 2013 and to eradicate the disease from livestock and wildlife. The West Taupo area, in the central North Island of New Zealand, was chronically infected with Tb in both domestic livestock herds (cattle and deer) and within wildlife populations (brushtail possum, ferret, feral deer and pigs). Through the development and implementation of a technically innovative management plan, this area is now approaching Tb free status. The case study / management plan reported here discusses the operational techniques and strategies that were implemented to achieve Tb clearance in the livestock herds and the possibilities of eradication from wildlife species. It particularly identifies the variations in control strategies that are required as population densities reduce and the challenges of maintaining strong effective control at low densities of some wildlife species, whilst not needing to control other species that were initially clinically diagnosed with Tb control. Use of diagnostic tools and education as an area moves through the cycle towards Tb freedom are as essential as the physical control activities. The use of intensive monitoring of both livestock and wildlife species as trend and performance indicators and the need to educate farmers, hunters and other land use groups become increasingly important.