Ultrastructural Studies of Acrosomal Formation in the Testis of Male Greater Cane Rat (Thryonomys swinderianus).

PURPOSE: In furthering the understanding of the process of spermatogenesis in the greater cane rat, this study describes the ultrastructural spermiogenic transformation and acrosomal formation in the testes of this hystricomorphic rodent that is currently undergoing domestication in parts of West Africa. MATERIALS AND METHODS: Testicular samples were obtained from ten sexually mature cane rats that were perfused-fixed using Karnovsky's fixative (phosphate buffered 2% paraformaldehyde - 2.5% glutaraldehyde fixative at pH 7.4). The samples were processed for ultrastructural analysis and examined under the transmission electron microscope. RESULTS: The testes of the cane rat showed uniqueness in its cellular associations and the ultrastructure of the spermatogenic cells especially in the formation of the acrosome. The spermatid differentiation and acrosomal formation occurred in 12 steps with the first three steps being the Golgi phase and the next three steps making up the cap phase. While the three steps that follow constitute the acrosomal phase, the last 3 steps make up the maturation phase. At the cap and acrosomal phases, the entire acrosomal system comprising the vesicle and granule covers the head of the spermatids with no clear indentation of the nuclear surface by the formed acrosome. Furthermore, elongated spermatids at the maturation phase contained abundance of nuclear vacuoles. CONCLUSION: This work has not only provided information that will further the understanding of spermatogenesis but also aid the understanding of acrosomal reaction in the reproduction of the greater cane rat.

Immunolocalization And Distribution Pattern of Estrogen (ERα and ERß) and Progesterone (PR) Receptors along the Excurrent Duct of Male Greater Cane Rat (Thryonomys swinderianus).

The excurrent duct, which plays vital roles in the reproductive biology of all male mammals, shows some structural variations among different species. Some hormones such as testosterone, estrogen and progesterone, through their different receptors, have been known to be involved in the normal functioning of the excurrent duct. Here we evaluated the presence, localization and patterns of distribution of three hormone receptors, estrogen alpha (ERα), estrogen beta (ERß) receptors and progesterone receptors (PR) along the excurrent duct of sexually matured male greater cane rats. Immunohistochemistry revealed presence of ERα in epididymal stroma but not epithelium, selective ERß staining in narrow & apical cells as well as unique presence of PR in caudal epididymis, which to the best of our knowledge, is the first report on the cellular localization of progesterone receptor in the cauda epididymis. The result suggests the possible involvement of not only estrogen but also progesterone in the modulation of epididymal function in greater cane rat.

Gross, histological and ultrastructural features of the bulbourethral gland in the greater cane rat (Thryonomys swinderianus).

The present study examines the structure and ultrastructure of the bulbourethral glands in 10 sexually matured male greater cane rats raised in captivity. Following anaesthesia, the rats were perfusion-fixed transcardially and the bulbourethral glands dissected out. Upon morphologic and morphometric analysis, the Cowper's glands were observed to have an average volume of 0.24±0.08 ml, a diameter of 6.3±0.6 mm and weighs 0.199±0.06 g. The paired, gourd-shaped tubuloalveolar glands were surrounded by dense connective tissues and separated into lobules by capsular septae. Each lobule consists of endpiece/secretory units and excretory ducts lined by simple glandular epithelium and pseudo-stratified epithelium, respectively. The round end pieces consisted of 8-10 pyramidal to columnar epithelial cells with flattened, basally located nuclei and granule-filled cytoplasm that bounded a narrow glandular lumen. The striking ultrastructural features of these secretory cells were the presence of some granules with uniform electron density and those with regions of lesser density as well as the absence of secretory vacuoles. Another unique characteristic of these secretory granules is the presence of electron dense strands radiating from their surfaces. The apical surfaces of the cells were also studded with abundant microvilli. From the findings, the structure of bulbourethral glands in the greater cane rat shows more resemblances to that of humans than to its rodent phylogeny. These findings serve as additional knowledge in the structural interpretation of the bulbourethral gland and its secretory products.

Chlamydia Infection; Plasma Peroxidation and Obesity in Tubal Infertility

Background: Genital tract infections and obesity are both sources of oxidative stress. Alterations in immune and antioxidant parameters may arise from this or from an indeterminate autoimmune mechanism. Objective: This study aimed to investigate the association of Chlamydial infection; obesity and oxidative response with tubal infertility in Nigerian women. Methods: It was a case-control study of 40 women with tubal infertility and 32 fertile women; respectively; recruited from the Infertility and Family Planning Clinics respectively; of the University College Hospital; Ibadan; Nigeria. Anthropometric indices were measured in each subject and endocervical swabs were taken to screen for current genital tract infection. Antioxidant; hormonal and immunologic analysis were performed on serum. Results: None of the subjects had current genital tract infections. Chlamydia trachomatis IgG positivity was significantly higher in infertile than in fertile subjects [OR 4.33; 95CI (0.078-0.681)]. No significant variations were observed in the anthropometric indices; antioxidant parameters and hormones between infertile and the fertile women. Body mass index correlated positively with oxidative stress in infertile subjects. Waist and hip circumferences correlated negatively with oestradiol in women with tubal infertility. Conclusion: Chlamydial infection is associated with tubal factor infertility; however; obesity seems to increase oxidative stress and reduce fertility potential in women with tubal factor infertility

The Gut and Lung Morphometry in Experimental and Natural Lineage 1 Variant of Peste des Petits Ruminants Virus Infection in Nigerian Goats / Morfometría Intestinal y Pulmonar en la Infección Natural y Experimental del Virus Peste des petits ruminants del linaje 1 en Cabras de Nigeria

The lung and gut morphometry in both natural and experimental Peste de petit ruminant (PPR) virus which are scanty in literature hence the need for this study. The goats that were submitted for necropsy in the Department of Veterinary Pathology University of Ibadan between 2009 and 2010 and the gross pathological diagnosis were PPR were enrolled in this study. The degree of pneumonia as a percentage of the total lung volume was estimated using standard methods. The gut morphometry of goats experimentally infected with PPR virus was also used. Student "T" test was used for the test of significance in evaluating the effect of age, sex and the lung consolidation pattern in natural PPR and analysis of the gut morphometry. Complicated PPR had significant higher pulmonary consolidation when compared with the uncomplicated PPR (p< 0.05). The pulmonary consolidation was significantly higher on the right lung with a mean percentage value of 6.54 than the left lung (p< 0.05). The caudal lobe was more consolidated than the cranial and middle lobes in natural PPR. The pulmonary consolidation was more in goats less than a year, while the buck had a significantly higher pulmonary consolidation than the does (p< 0.05). There was no significant difference in the mean length of the villi and width of the villi of PPR virus infected goats when compared to the control, however a significant difference was observed in the cryptal depth (p< 0.05). There was a significant difference in the mean villi length and cryptal depth of goats with complicated PPR (Mannheimia hemolytica) infected goats (p< 0.05) relative to the control. From this study, it showed that most natural PPR were complicated with bacteria and this complication may have contributed to the fatality associated with PPR especially those caused by lineage 1 viruses. This study also showed that secondary bacterial involvement in course of PPR affect the gut morphometry and that could account for the severity of intestinal lesion commonly observed with field PPR in Nigerian goats.

La morfometría del pulmón y el intestino en la infección del virus Peste des petits ruminants (PRR) de forma natural así como experimental es escaza en la literatura, de ahí la necesidad de este estudio. Fueron incluidas en este estudio las cabras que fueron sometidas a autopsia en el Departamento de Patología Veterinaria de la Universidad de Ibadan entre 2009 y 2010, con diagnóstico patológico macroscópico de PPR. El grado de neumonía como porcentaje del volumen pulmonar total fue estimado mediante los métodos estándar. También fue determinada la morfometría del intestino de las cabras infectadas experimentalmente con el virus PPR. Se utilizó la prueba "T" de Student para determinar la significancia en la evaluación de los efectos de edad, sexo, patrón de consolidación pulmonar en PPR natural y análisis de la morfometría intestinal. La PPR complicada tuvo una consolidación pulmonar altamente significativa en comparación con la PPR no complicada (p <0,05). La consolidación pulmonar fue significativamente mayor en el pulmón derecho, con un valor porcentaje promedio de 6,54 en comparación al pulmón izquierdo (p <0,05). El lóbulo caudal fue más consolidado que los lóbulos craneal y medio en presencia del PPR natural. La consolidación pulmonar fue más frecuente en caprinos menores de un año, mientras que los machos cabríos tuvieron una consolidación pulmonar significativamente más alta (p <0,05). No hubo diferencias significativas en la longitud y ancho promedio de las vellosidades en cabras infectadas con PPR en comparación con el control, pero se observó una diferencia significativa en la profundidad de las criptas (p <0,05). Hubo diferencia significativa en la longitud de las vellosidades y la profundidad media de las criptas en las cabras infectadas con PPR complicada (Mannheimia haemolytica) (p <0,05) en relación al control. A partir de este estudio, se demostró que las infecciones con PPR natural se complicaron con bacterias, y estas complicaciones pueden haber contribuido a la mortalidad asociada el PPR, especialmente las causadas por el virus del linaje 1. Este estudio también mostró que la participación bacteriana secundaria en el curso de la PPR afecta la morfometría intestinal y que podría dar cuenta de la gravedad de la lesión intestinal observada comúnmente en la infección de PPR en cabras de Nigeria.

Chlamydial infection, plasma peroxidation and obesity in tubal infertility.

BACKGROUND: Genital tract infections and obesity are both sources of oxidative stress. Alterations in immune and antioxidant parameters may arise from this or from an indeterminate autoimmune mechanism. OBJECTIVE: This study aimed to investigate the association of Chlamydial infection, obesity and oxidative response with tubal infertility in Nigerian women. METHODS: It was a case-control study of 40 women with tubal infertility and 32 fertile women, respectively, recruited from the Infertility and Family Planning Clinics respectively, of the University College Hospital, Ibadan, Nigeria. Anthropometric indices were measured in each subject and endocervical swabs were taken to screen for current genital tract infection. Antioxidant, hormonal and immunologic analysis were performed on serum. RESULTS: None of the subjects had current genital tract infections. Chlamydia trachomatis IgG positivity was significantly higher in infertile than in fertile subjects [OR 4.33; 95%CI (0.078-0.681)]. No significant variations were observed in the anthropometric indices, antioxidant parameters and hormones between infertile and the fertile women. Body mass index correlated positively with oxidative stress in infertile subjects. Waist and hip circumferences correlated negatively with oestradiol in women with tubal infertility. CONCLUSION: Chlamydial infection is associated with tubal factor infertility, however, obesity seems to increase oxidative stress and reduce fertility potential in women with tubal factor infertility.

Effects of melarsamine hydrochloride (Cymelarsan) and diminazene aceturate (Berenil) on the pathology of experimental Trypanosoma brucei infection in red fronted gazelles (Gazella rufifrons).

An experimental infection of red fronted gazelles (Gazella rufifrons) with Trypanosoma brucei strain MKAR/84/NITR/6 was carried out. Two waves of parasitaemia which corresponded with a significant decline (p<0.05) in packed cell volume (PCV) was encountered in the infected untreated controls and those treated at day 8 post-infection with a sub-optimal dosage of diminazene aceturate (Berenil) at 3.5 mg/kg body weight. At postmortem, hepatomegally, splenomegally, lymphadenopathy, nephritis, myocardial degeneration with pulmonary oedema was observed in the two groups. Similarly, histopathological studies of some organs revealed interstitial haemorrhages, severe degenerative changes with cellular infiltrations. On the other hand, those treated by day 8 post-infection with melarsamine hydrochloride (Cymelarsan) at 0.3 mg/kg, 0.6 mg/kg or diminazene aceturate (Berenil) at 7.0 mg/kg body weight had apparently normal organs at the end of the experiment. These results suggest that, T. brucei can cause severe pathological changes in untreated red fronted gazelles (Gazella rufifrons). However, treatments at the onset of parasitaemia, by day 8 post-infection with melarsamine hydrochloride (Cymelarsan) at 0.3 and 0.6 mg/kg or diminazene aceturate (Berenil) at 7.0 mg/kg body weight ameliorated the deleterious effects of the infection in the gazelles.

Anti-oxidant and anti-hyperglycemic activities of musa sapientum root extracts in alloxan-induced diabetic rats.

Anti-hyperglycemic and anti-oxidant properties of methanolic (MEMS) and aqueous (AEMS) extracts of Musa sapientum roots were investigated in alloxan-induced diabetic rats. Thirty adult male Wistar albino rats divided into five groups of 6 rats each were used: group 1--non-diabetic untreated (controls), group 2--diabetic untreated, and groups 3, 4 and 5--diabetic rats treated with 250 mg/kg bodyweight MEMS and AEMS, and 500 mg/kg bodyweight glibenclamide (a standard anti-diabetic drug), respectively. There was severe progressive weight loss in the untreated diabetic rats, while the rats in all the treated diabetic groups gained weight. While there was progressive hyperglycaemia in untreated diabetic rats; with blood glucose levels reaching a peak of 335.5 +/- 1.1 mg/dl on day 7 post-induction, compared to 76.8 +/- 0.8 mg/dl on day 0, these values were reduced to 80.7 +/- 0.5, 86.6 +/- 0.6 and 86.8 +/- 0.5 in MEMS, AEMS and glibenclamide-treated diabetic rats 15 days post-treatment. Also there were decreases in serum lipid peroxidation and increases in serum superoxide dismutase activities in MEMS, AEMS and glibenclamide-treated diabetic rats 15 days post-treatment. Lesions observed in the organs of untreated diabetic rats include selective necrosis of pancreatic beta islet cells, hepatocellular degeneration and necrosis, glomerulonephrosis and cardiovascular degeneration. Treatment of diabetic rats with AEMS and glibenclamide caused a total mitigation, while treatment with the MEMS achieved partial but considerable reduction in the severity of the lesions. It is concluded that aqueous and methanolic extracts of Musa sapientum roots possess anti-diabetic activities comparable to glibenclamide.

Histopathological studies on the effects of the ethanolic extract of the fruits of Garcinia kola on selected organs of the dog

The ethanolic extract of Garcinia kola was administered to dogs to investigate the possible effects on selected organs of the dog. Two doses of the extract (500mg/Kg and lOOOmg/Kg) were daily administered to the test animals for a period of 6 weeks. A dose related response was observed in the severity of histopathological changes observed in the testes, liver, kidney and small intestine of animals in the test groups. Despite the reported potentially beneficial effects of Garcinia kola, its use as a medicinal plant should be with great caution.

Fue administrado a perros extracto etanólico de Garcinia kola, para investigar los posibles efectos sobre determinados órganos. Dos dosis del extracto (500mg/Kg y lOOOmg/Kg) fueron administradas diariamente a los animales durante un periodo de 6 semanas. Se observó una relación de la dosis con la gravedad en la respuesta de los cambios histopatológicos observados en testículos, hígado, riñón e intestino delgado de los animales. A pesar de los informes sobre efectos potencialmente beneficiosos de Garcinia kola, su uso como planta medicinal debe ser con mucha precaución.

Non-immune control of trypanosomosis: in vitro oxidative burst of PMA- and trypanosome-stimulated neutrophils of Boran and N'Dama cattle.

An in vitro assay that measures the generation of superoxide anions (O2-) was used to assess the level of oxidative burst of phorbol myristate acetate (PMA)- and trypanosome-stimulated neutrophils isolated from healthy Boran and N'Dama cattle, and those infected with Trypanosoma congolense. PMA stimulation of healthy bovine neutrophils resulted in between 300-400 % increase in O2- generation. Neutrophils of Boran cattle exhibited slightly higher but insignificant O2- generation capacity than those of the N'Dama breed. In vitro stimulation by trypanosomes of neutrophils isolated from Trypanosoma congolense-infected cattle caused significant increases in O2- generation, especially on days 14, 28 and 42 post-infection, of both breeds of cattle. No significant differences were observed in O2- generation capacity of the neutrophils of both breeds of infected cattle throughout the period of assay. The results of this study have shown that PMA and trypanosomes do cause an enhanced in vitro oxidative burst, hence trypanosome phagocytosis and killing activity of neutrophils. Neutrophils have been shown to play very significant roles in parasite clearance, hence reduction of trypanosome parasitaemia. The rates of both in vitro generation of O2- and trypanosome phagocytosis over time did not differ significantly between Boran and N'Dama breeds of cattle, even during T congolense infection in this study. Hence, it may be inferred that sustained and higher parasitaemia, more pronounced neutropenia, inadequate bone marrow response and less effective trypanosome-specific immune response, rather than defective neutrophil trypanosome destruction, may be the problem of trypanosusceptible cattle breeds.

Pathobiochemical mechanisms involved in the control of the disease caused by Trypanosoma congolense in African grey duiker (Sylvicapra grimmia).

The course of Trypanosoma congolense infections in African grey duiker (Sylvicapra grimmia) and sheep and goats were studied. Several parameters suggested that the grey duiker was much more resistant to trypanosomosis than sheep and goats. They showed increases in weight during infection, had a much longer pre-patent period, and their peak parasitaemia levels were about 100-fold lower than those of sheep and goats. Parasites were no longer detected in grey duiker blood 35 days after infection. Anaemia, measured as drops in packed cell volume (PCV), haemoglobin (Hb) concentration and erythrocyte (RBC) counts were not observed in the grey duiker. In contrast, sheep and goats suffered severe weight losses and had continuously high parasitaemia levels. Sheep and goats developed progressively severe normocytic normochromic anaemia and leucopenia from day 14 post-infection onwards. Serum levels of total protein, globulin and albumin of grey duiker did not change significantly throughout the course of infection, while the levels of total serum protein, globulin and gamma-globulin exhibited significant increases from day 21 post-infection onwards in sheep and goats, with peak values recorded on 28 and 35 days post-infection in sheep and goats, respectively. There were inconsistent variations in albumin levels in sheep and goats throughout the course of infection. There were no significant changes in erythrocyte activities of AST and ALT, while there were transient but significant elevations of ALP level on day 35, and GGT levels between 14 and 35 days post-infection in grey duiker. Conversely, the levels of all the enzymes were progressively depressed, especially from 14 to 49 days post-infection. In vitro erythrocyte peroxidation remained relatively unchanged throughout the period of the experiment in the grey duiker, except for slight but significant increase on day 42 post-infection. However, in vitro erythrocyte peroxidation increased significantly by between 100 and 300% of pre-infection levels from 14th to 42nd day p.i. both in sheep and goats, before returning to pre-infection levels after 14 days of treatment. Haematological values, serum and erythrocyte indices studied returned to near pre-infection levels 14 days after treatment with Berenil((R)). It is concluded that the grey duiker is inherently trypanotolerant. This is shown by its ability to control parasitaemia, suffer less severe anaemia, and to a relative degree resist pathobiochemical derangements of some serum and erythrocyte metabolites and enzymes, as well as reduction of infection-induced erythrocyte lipid peroxidase damage than sheep and goats.

Serological survey of salmonellosis in grey duiker (Sylvicapra grimmia) in Asejire, Irewole Local Government Area, Osun State, Nigeria.

A serological survey of salmonellosis in grey duiker was carried out in Asejire, Irewole Local Government Area (LGA), Osun State, between August and September, 1996. The sera of 50 animals were screened for antibodies to salmonella flagella (H) and/or somatic (O) antigens. Ten "bushmeat" hawkers/processors were interviewed on their state of health. Twenty-two (44%) and 3 (6%) had antibodies to the H and O antigens, respectively. Nine (18%) had antibodies to both the H and O antigens. Antibody titres of 1/320 were detected against Salmonella paratyphi serotypes B (12%) and C (2%) and S. typhi serotype D (8%). Four (40%) of the interviewed hawkers/processors had had 'fever' in the recent past, and 80% of them usually take traditional herbal medicines for most fever-related ailments. A human typhoid epidemiological surveillance carried out within the LGA between 15th March and 4th April, 1995 revealed 64 cases of tentatively diagnosed salmonellosis. Eleven (17.2%) cases were conclusively diagnosed as typhoid fever by laboratory examination. High antibody titres against H and O antigens of S. paratyphi serotype B and S. typhi serotype were detected. Two (3.1%) of the patients died. The public health importance of these findings is discussed.

In vitro erythrophagocytosis by cultured macrophages stimulated with extraneous substances and those isolated from the blood, spleen and bone marrow of Boran and N'Dama cattle infected with Trypanosoma congolense and Trypanosoma vivax.

A standard radioactive chromium (51Cr) release assay was used to assess the in vitro phagocytosis and lysis of bovine erythrocytes by cultured splenic, bone marrow and peripheral blood monocyte-derived (PBM) macrophages isolated from healthy and Trypanosoma congolense and T. vivax-infected cattle of the Boran and N'Dama breeds. Recombinant cytokines (rHuTNF-alpha and rBolFN-gamma) and non-acid-dialysed peripheral blood mononuclear cell (PBMNC) culture supernatants stimulated these PBM for enhanced activities. The stimulants caused increases in the rate of erythrocyte phagocytosis and lysis by cultured PBM in a concentration-dependent manner. But very high stimulant concentrations caused deceased in vitro erythrophagocytosis. However, bacterial lipopolysaccharide (LPS) and acid-dialysed PBMNC culture supernatants did not cause any increase in cultured PBM erythrophagocytosis. In vitro erythrocyte phagocytosis and lysis by splenic, bone marrow and peripheral blood monocyte (PBM)-derived macrophages of Boran breed of cattle infected with Trypanosoma congolense increased from 14 days post-infection (DPI) onwards and thereafter maintained at various levels above pre-infection. Cultured splenic macrophages showed the greatest erythrocyte destruction capability while PBM-derived macrophages was the least. The rates of in vitro erythrocyte phagocytosis and lysis were higher with the cultured PBM of the Boran than those of the N'Dama cattle during T. congolense infection. The rate of in vitro erythrocyte destruction was however, similar in both groups of cattle during T. vivax infection. These results correlated positively with the dynamics and degree of anaemia developed by these groups of animals during both T. congolense and T. vivax infections. Cattle infected with T. congolense and T. vivax developed varying degrees of normocytic normochromic anaemia during infection. Boran cattle developed a more severe anaemia, and had to be treated with diminazine aceturate, than N'Dama cattle during T. congolense infection. Both breeds of cattle developed a milder but similar degree of anaemia during T. vivax infection. None of the animals were treated. The results of this study indicated a role of in vivo macrophage stimulatory factors, notably cytokines such as TNF-alpha and IFN-gamma in host's serum, as well as parasite antigens, which may act singly or in concert, in the process of enhanced erythrocyte destruction, hence anaemia by the mononuclear phagocytic system (MPS) during bovine trypanosomosis.

Differential expression of surface membrane antigens on bovine monocytes activated with recombinant cytokines and during Trypanosoma congolense infection.

The expression of surface membrane antigens on peripheral blood monocytes (PBM) of cattle of the Boran and N'Dama breeds activated with recombinant cytokines (TNF-alpha and IFN-gamma) and during experimental infection with Trypanosoma congolense was investigated using monoclonal antibodies (MoAbs) and fluorescein-activated cell sorter (FACS). The surface antigens investigated were C3bi receptor, major histocompartibility (MHC) II complex (Ia antigen) and two monocyte/macrophage (Mphi) differentiation antigens. The study revealed that both cytokines caused the enhancement of the expression of all the PBM surface antigens studied. rBolFN-gammaat low concentrations was more efficient in causing the activation of PBM. While the PBM of Boran cattle were more significantly activated to express the C3bi receptor vis-à-vis the Ia antigen than N'Dama cattle, the reverse was the case with the PBM of N'Dama cattle which expressed more Ia antigens than Boran PBM. Similar results were observed during T. congolense infection in the two breeds of cattle. The significantly higher expression of C3bi receptor and correspondingly lower Ia antigen expression by the PBM of Boran cattle, both during trypanosomosis and in vitro may be responsible for the higher rate of erythrocyte phagocytosis, hence the development of more severe anaemia by Boran cattle during trypanosomosis than N'Dama. In addition, the expression of significantly higher numbers of Ia antigen by N'Dama Mphi, hence are more able to process, present and initiate better trypanosome antigen-specific immune response than Boran cattle during infection. These two attributes are known genetic characteristics of trypanotolerance in cattle.

Role of the chancre in induction of immunity to tsetse-transmitted Trypanosoma (Nannomonas) congolense in goats.

Local skin reactions (chancres) developed in goats at the sites of deposition, by tsetse flies, of metacyclics of Trypanosoma congolense. The chancres developed much faster and were more pronounced when ten infected tsetse were allowed to feed on a spot as compared to only one fly per spot. The initial host cellular reaction in the chancre was predominantly polymorphonuclear, followed at the peak of development of the chancre by a predominantly lymphoblastic and plasmacytic reaction. Trypanosomes were found in various stages of division as well as degeneration in chancre biopsies taken at various days post-infection (p.i.). Most of the trypanosomes recovered from the chancre tissue fluid were found to bear the same variable surface glycoprotein (VSG) epitopes as the corresponding metacyclics for as long as 13 days p.i., as revealed by indirect immunofluorescence using mouse anti-metacyclic VSG hyperimmune sera and monoclonal antibodies. Immunization of goats with metacyclic trypanosomes, by exposure to infected tsetse bites followed by treatment of the infected goats on day 13 p.i., gave rise to the development of protection to homologous tsetse-transmitted challenge, whilst immunization by intravenous inoculation of the metacyclics did not induce such protection. Chancre formation would thus appear to be vital for the induction of comprehensive immune recognition of the metacyclic variable antigen repertoire deposited in the skin by infected tsetse, and hence development of protective immunity.