Molecular characterization and phylogenetic analysis of Marek's disease virus in chickens from Ogun State, Nigeria.

Marek's disease (MD) is caused by oncogenic MD virus serotype 1 (MDV1) and is characterized by lymphoproliferative lesions resulting in high morbidity and mortality in chickens. Despite being ubiquitous on poultry farms, there is a dearth of information on its molecular characteristics in Nigeria. This study aimed at characterizing three virulence genes (Meq, pp38, and vIL-8) of MDV1 from chickens in Ogun state, Nigeria. Blood, feather quill, and tumour samples of chickens from different commercial poultry farms in Ogun State were pooled, spotted on 107 FTA cards, and screened for MDV1 by polymerase chain reaction (PCR). Phylogenetic analysis was carried out to compare Nigerian MDV1 Meq, pp38, and vIL-8 genes sequences with the published references. Thirteen samples were MDV1-positive and the Meq, as well as pp38, and vIL-8 genes from the different samples were 100% identical. The Meq genes contained 339 amino acids (aa) with three PPPP motifs in the transactivation domain and two interruptions of the PPPP motifs due to proline-to-arginine substitutions at positions 176 and 217 resulting in a 20.88% proline composition. Phylogenetic analysis revealed that the Meq gene clustered with strains from Egypt and very virulent ATE2539 strain from Hungary. Mutations were observed in the pp38 protein (at positions 107 and 109) and vIL-8 protein (at positions 4 and 31). Based on the molecular analysis of the three genes, the results indicate the presence of MDV1 with virulence signatures; therefore, further studies on in vivo pathotyping of Nigerian MDV1 from all states should be performed.RESEARCH HIGHLIGHTS Meq, pp38 and vIL-8 genes were 100% identical between Nigerian MDV strains.Proline content in Nigerian meq gene was 20.88% with two PPPP motifs interruptions.Meq, pp38 and vIL-8 genes of Nigerian MDV were similar to Egyptian and Indian strains.

Erythrocyte Sedimentation Rates and Leukogram Changes in Canine Model of Osteoarthritis.

Inflammatory markers such as erythrocyte sedimentation rates (ESR) have been evaluated in humans withosteoarthritis (OA). However, there has been no record of evaluation of ESR during OA in dogs. Changes in erythrocytesedimentation rates (ESR) and leukogram functions were evaluated following experimental knee osteoarthritis (OA).Tendogs of both sexes with (mean weight = 12.4 ± 1.8kg) were used. Experimental OA was induced in the right knee, using thegroove model and confirmed radiographically using evidence of joint space narrowing and presence of osteophytes. Gaitwas assessed subjectively and scores (GAS) were assigned. Blood was obtained fortnightly for the determination of ESR,total white blood cell (tWBC), neutrophil and lymphocyte counts, while knee radiographs were obtained fortnightly fortwelve weeks. Radiographic scores (RAS), GAS, ESR and leukocyte parameters between the different time points werecompared with ANOVA. Correlation between parameters was evaluated using Pearson's correlation. A "P" value less than0.05 was considered significant. Both ESR and neutrophil/lymphocyte (N/L) ratio increased from week 0 to week 12 of OA.However, tWBC, neutrophil and lymphocyte counts did not differ significantly. Both GAS and RAS increased up to week 4and 6 of OA respectively. Erythrocyte sedimentation rates was significantly (p= 0.033) and positively correlated (r=0.793)with N/L ratio, but negatively and slightly correlated (r= -0.843) with GAS. There was no significant correlation betweenESR and RAS. It was concluded that both ESR and N/L ratio might be useful in monitoring progression of OA in dogs.

Potentially zoonotic shiga toxin-producing Escherichia coli serogroups in the faeces and meat of food-producing animals in Ibadan, Nigeria.

Shiga toxin-producing Escherichia coli (STEC) are major food-borne pathogens associated with gastroenteritis and sometimes fatal haemolytic uraemic syndrome complication. Farm animals are asymptomatic carriers of STEC and contaminated meat is an important vehicle for zoonotic transmission from animals to humans. This study investigated the presence, virulence traits and antimicrobial susceptibility of seven potentially human pathogenic STEC serogroups (O157, O26, O91, O103, O111, O128 and O145) in the faeces and meat of food-producing animals in Ibadan, Nigeria. One hundred and fifty-four (7.3%) of 2133 samples were positive for STEC serogroups. The pathogens were detected in the faeces of cattle (15.2%), sheep (10.7%), goats (7.5%) and pigs (5.6%) as well as in beef (3.8%), goat-meat (1.7%) and pork (4.0%). All seven investigated STEC serogroups were found in cattle, all except O145 were found in sheep, three serogroups (O157, O26 and O111) were found in goats and three (O157, O111 and O128) in pigs. The rate of detection of each of the serogroups in all 2133 samples was: O157 (5.0%), O26 (0.2%), O91 (0.3%), O103 (0.3%), O111 (1.0%), O128 (0.2%) and O145 (0.1%). Of all 154 isolates, 11.0% had shiga toxin type 1 gene (stx(1)), 25.3% had stx(2) and 41.6% had stx(1)/stx(2); intimin gene (eaeA) was detected in 56.5% and enterohaemolysin gene (hlyA) in 75.3%. Among the O157 isolates, 24.5% were negative for stx genes but positive for eaeA and/or hlyA while 7.6% were negative for all four virulence genes. Fourteen different combinations of virulence genes were encountered but stx(1)/stx(2)/eaeA/hlyA combination was the most predominant. The percentage resistance of the isolates to the tested antimicrobial agents was: ampicillin (82.5%), chloramphenicol (42.9%), ciprofloxacin (22.1%), enrofloxacin (25.3%), nalidixic acid (37.7%), neomycin (24.0%), norfloxacin (20.8%), streptomycin (50.7%) and tetracycline (75.3%). One hundred and forty-eight (96.1%) of all 154 isolates were resistant to at least one of the tested antimicrobial agents while 69.5% were categorised as multi-drug resistant. Potentially pathogenic multi-drug resistant STEC isolates were recovered from the meat production chain in Nigeria. Unhygienic practices that predominate during slaughter and processing were observed to have contributed to faecal contamination and presence of STEC in meat.

An outbreak of African Swine Fever in Nigeria: virus isolation and molecular characterization of the VP72 gene of a first isolate from West Africa.

The isolation of 98/ASF/NG, a strain of African Swine Fever Virus (ASFV) associated with a 1998 epizootic in Nigeria, is reported. This first isolate of the virus from West Africa was identified through a successful polymerase chain reaction (PCR) amplification and sequencing of a 280 base pair (bp) fragment of the Major Capsid Protein (VP72) gene. Further amplification and sequence analysis of a 1.9 kilobase pair (kbp) fragment encompassing the complete VP72 gene showed that the isolate has a 92.2%, 92.4%, and 97.2% homology with previously sequenced Ugandan, Dominican Republican and Spanish isolates respectively. Of the 50 nucleotide changes observed in this highly conserved gene, 45 were found to result in 40 amino acid changes clustered around the central region (position 426 to 516) of the VP 72 protein while changes at the remaining 5 positions were silent. These changes also led to the loss of two out of the seven potential N-glycosylation sites which are in this gene conserved among all isolates. The possible epizootiological implications of such mutations in a highly conserved gene of a DNA virus is discussed in relation to this outbreak.

Immune lysis of Trypanosoma congolense: generation of a soluble covalent complex of variant surface glycoprotein and bovine complement component C3b.

Organisms of Trypanosoma congolense variant antigenic type TC13 (VAT TC13) were incubated, at 37 degrees C for 60 min, with fresh bovine serum in the presence of antibody specific for the variant surface glycoprotein (VSG). Upon immune lysis, soluble VSG (54 kDa) and a larger complex (about 225 kDa), containing VSG, was detected in the supernatant fluid of the mixture. Neither soluble VSG nor the VSG complex were detected when fresh bovine serum was incubated with organisms of T. congolense in the absence of specific antibody. Within a narrow range of low antibody concentration, the release of soluble VSG and the formation of the VSG complex were correlated with the amount of specific antibody added to the mixture. The VSG complex could be precipitated with rabbit antibodies specific for VSG of VAT TC13 or antibodies specific for bovine complement C3. The VSG complex was detected by Western blot with rabbit anti-VSG of VAT TC13 as well as rabbit antibovine C3. The complex was found to consist of VSG covalently bound to bovine complement component C3b. Potential pathophysiological implications are discussed.

Susceptibility of Nigerian west African dwarf and red Sokoto goats to a strain of Trypanosoma congolense.

West African Dwarf (WAD) and Red Sokoto (RS) goats were experimentally infected with the Kafanchan strain of Trypanosoma congolense and the course of the infection was monitored. The organism was pathogenic and produced fatal disease in the goats, which was characterized by rapid progressive anaemia, leucocytosis, weight loss and death. All RS goats died within 11 days of infection, and had a mean reduction in packed cell volume (PCV) of 11%. In West African Dwarf goats, one death occurred on Day 13 post-infection with a mean drop in PCV of 9%. Statistically significant (P < 0.05) mean reductions in values of PCV, haemoglobin and red blood cell counts were observed between the infected and control animals of both breeds, and also between the infected WAD and infected RS goats. The anaemia produced was macrocytic. Leucocytosis characterized by neutropenia and lymphocytosis was observed among infected WAD goats, but leucopenia characterized by neutrophilia and lymphopenia was observed in infected RS goats. Infected WAD goats recorded some positive unit weight gain in spite of the infection. It was concluded that the RS breed of goats is more susceptible to T. congolense infection than the WAD breed.

The response of pigs experimentally infected with Trypanosoma brucei to isometamidium chloride therapy and the relation to nutrition.

Growing pigs were placed on feeds with high (Group A), medium (B) and low (C) dietary energy and were infected with a virulent stock of T. brucei. Eight weeks later, the infected pigs were treated with isometamidium chloride at 1 mg/kg live weight and all pigs were subsequently placed on a high energy diet to investigate their response to therapy. Clearance of T. brucei from blood was completed 72h after treatment. There was no evidence of relapsed infection up to eight weeks after treatment. Red blood cell parameters returned to normal four to six weeks after treatment with responses being fastest in Group A, B and C had gained about two-thirds of the live weight gains of their non-infected pair-fed controls. It appears that the retarded weight gain as a result of the infection persisted after therapy since drug-treated pigs did not gain as much weight as their non-infected controls.

Changes in levels of transaminases in goats experimentally infected with Trypanosoma congolense.

Goats were experimentally infected with Trypanosoma congolense and then treated with Berenil after 9 days of infection. The infection produced increases in glutamate oxalacetate transaminase (GOT) and glutamate pyruvic transaminase (GPT) values. Mean GOT values in infected West African dwarf goats were generally lower than in infected Red Sokoto goats. Treatment with Berenil did not produce any significant effect on their levels probably because of the relapse infection recorded in this study.

Plasma levels of proteins of the alternative complement pathway in inbred mice that differ in resistance to Trypanosoma congolense infections.

Inbred BALB/c, A/J, and C57B1/6J mice were infected with Trypanosoma congolense (Trans Mara strain), clone TC13, and monitored for parasitemia, survival times, and plasma levels of complement components C3, C5, factor B, and factor H. Parasitemia was highest in BALB/c, intermediate in A/J, and lowest in C57Bl/6J mice. The mean survival times were 11.5 +/- 0.9, 23.8 +/- 2.3, and 119 +/- 26 days for BALB/c, A/J, and C57Bl/6J mice, respectively. Preinfection levels of factor H were significantly correlated with survival times (r = 0.7722, P less than 0.001). Marked differences were observed between the plasma levels of C3, factor B, and factor H in the 3 mouse strains following infection. Complement C5 levels showed the fewest changes. In the initial postinfection period, BALB/c mice had highest increases in the levels of the 4 complement proteins but also had the greatest declines toward the end of the infection. Factor H levels showed a biphasic increase in BALB/c and C57Bl/6J, but not in A/J mice, with peaks at days 3 and 9. Complement C3 levels declined in all mice toward the terminal stage of the disease. In the late stages of infection, factor B levels markedly decreased in BALB/c but significantly increased in C57Bl/6J mice. Factor B levels measured at the terminal stages in BALB/c, A/J, and C57Bl/6J were correlated positively with their respective survival times (r = 0.714, P less than 0.01). The results suggest that genetic differences in the alternative complement pathway might affect the resistance to T. congolense infections.

The effect of Trypanosoma brucei infection on serum biochemical parameters in boars on different planes of dietary energy.

Young boars were placed on diets with either low or high dietary energy and subsequently infected with a virulent stock of Trypanosoma brucei. The effects of dietary energy level and infection on some serum biochemical parameters were evaluated up to 7 weeks post-infection (p.i.). There were no significant changes in serum electrolyte (Na+, K+) concentrations resulting from dietary energy level and/or the infection. Serum total protein and albumin levels significantly decreased in both groups of infected boars, the decline being greater in those on the low-energy diet. Infection was accompanied by a rise in serum transaminase (serum aspartate and alanine aminotransferases) levels which were higher in infected boars on the low-energy diet. The serum testosterone concentration declined in both groups of infected boars with the fall being more pronounced in the group on the low-energy diet. The results indicated that the reproductive efficiency of boars may be modulated by nutrition and that adequate feeding may assist in ameliorating the deleterious effects of trypanosomiasis on production in endemic areas.

Studies on west African dwarf sheep: incidence of perinatal mortality in Nigeria.

A study was conducted on the incidence of perinatal lamb mortality with extension up to 30 days of age (PMR 30) in West African Dwarf sheep in Ibadan, Nigeria, over a three year period. An overall PMR 30 of 19.9% was recorded. This consisted of an abortion rate of 3.0%, a still-birth rate of 2.7% and a neonatal lamb mortality rate of 15.6% up to 30 days of age (LMR). Abortion rate was significantly higher in pregnancies with multiple foetuses and during the dry sea son (P less than 0.01 and P less than 0.05 respectively). Conversely, the still-birth rate was slightly higher in single lambs (P less than 0.05). The LMR for males (17.1%) and that for females (13.4%) were not significantly (P greater than 0.05) different. Also the LMR for single-born (14.4%), twin-born (15.4%) and triplets (26.7%) did not significantly differ (P greater than 0.05). The optimum birth weight for survival of the breed appeared to be 2.6 to 3 kg. Regression analysis showed a 28.3% increase in birth weight and all live-born lambs with birth weights of 0.9 kg or less died during the neonatal period. About two-thirds (67.2%) of all neonatal deaths occurred during the first week of life. There was no significant effect of season on LMR. The perinatal mortality rates recorded among West African Dwarf sheep in this study do not seem to significantly differ from corresponding figures recorded in other parts of the world.

Pathogenicity of Trypanosoma brucei brucei in experimentally infected pigs.

An experimental infection of 4-to 5-month old pigs with a stock of Trypanosoma brucei brucei resulted in a high parasitaemia, anorexia, pyrexia and a decline in the packed cell volume by one third. Nervous sign of circling and wobbling of the hind legs occurred in one of the pigs which at necropsy revealed a very severe meningo-encephalitis and the presence of trypanosomes in the brain. These results confirm that T. b. brucei might cause a severe disease in pigs.

Survival and infectivity of Trypanosoma brucei in refrigerated pig blood.

Trypanosoma brucei parasites survived for 72 h or longer in refrigerated pig blood. The survival period was directly proportional to the initial parasite concentration of the sample. Infectivity of the parasites declined faster than survival, being less than 1 per 10(5) motile organisms at 72 h. The stage of infection in the pig (early vs. late) did not appear to influence subsequent survival periods or infectivity of the parasites in vitro.

The relationship between dietary energy levels and the severity of Trypanosoma brucei infection in growing pigs.

Growing pigs were placed on high, medium and low planes of dietary energy and were infected with a virulent strain of Trypanosoma brucei. During an 8-week period post-infection (p.i.), the respective liveweight gains by infected pigs on high, medium and low energy levels were 52.1, 21.2 and 38.5%, respectively, of the corresponding gains by non-infected control pigs. There was a fall in red blood cell values p.i. which worsened with decreasing energy levels. Leucocytosis was observed in all infected pig groups and was mainly due to lymphocytosis. By 6 weeks p.i., the lymphocyte count had returned to near normal values in pigs on high and medium energy levels, but was persistently high in those on a low energy level. Neutropaenia was observed in all infected pig groups and persisted until 8 weeks p.i. The results indicated that nutrition modulates the host response to infection with trypanosomes.

Experimental infection of Red Sokoto goats with Salmonella typhimurium.

Salmonella typhimurium infection was experimentally induced in goats by administering 2 x 10(10) organisms per os. The disease produced was characterized by pyrexia, diarrhoea and neutrophilia. One goat died from septicaemia. Somatic "0" agglutinins were detected 14 days post infection (p.i.). Excretion of organisms in faeces ceased by 6 weeks p.i. Goats which recovered from primary infection were refractory to a secondary challenge with 2 x 10(11) organisms. The results indicate that in the absence of signs of ill-health, the detection of neutrophilia and "0" agglutinins and isolation of Salmonella organisms from faeces mainly served as evidence of recent infection.

Enhanced resistance of highly susceptible Balb/c mice to infection with Trypanosoma congolense after infection and cure.

Balb/c and C57Bl/6 mice were cured with Berenil after infection with cloned organisms of Trypanosoma congolense and challenged with homologous or heterologous variants. The mice were fully protected against infection with 10(3) but not 10(5) organisms of the homologous variant. Normal Balb/c mice infected with 10(5) organisms developed uncontrolled parasitemia and had a mean survival time of 8.4 days. Challenge of drug-cured Balb/c mice with 10(5) organisms of the homologous variant established an infection associated with prolonged prepatent period, control of the first peak of parasitemia, and prolonged survival time (36 days). Indirect immunofluorescent and agglutination tests on live trypanosomes revealed that the "delayed" population of the first peak of parasitemia consisted of variants other than that used for challenge. No protection of drug-cured Balb/c mice was obtained following challenge with 10(1)-10(5) organisms of a heterologous variant. Passive transfer of variant-specific antiserum protected mice against infection with 10(3) organisms. Against infection with 10(5) organisms, it resulted in a prolonged prepatent period but had no effect on severity of parasitemia or duration of survival. There was no evidence for persistence of Berenil, which potentially could affect resistance. It was concluded that enhanced immunity in drug-cured Balb/c mice was due to (a) antibody to the variant surface glycoprotein and (b) another, yet unidentified, synergistically acting immune response to the parasite. Possible mechanisms are discussed.

Pathogenicity of Eperythrozoon suis alone and when mixed with Babesia trautmanni in experimentally-infected pigs.

The pathogenicity of a single infection with Eperythrozoon suis and of a mixed infection of E. suis and Babesia trautmanni in experimentally-infected pigs, was described. In the pigs with a single infection, parasitaemia of E. suis was observed 4 days after blood inoculation. Although parasitaemia with this parasite was also observed on the 4th day in a mixed infection, the parasitaemia of B. trautmanni was not noted until 14 days after inoculation at a period when that of E. suis had started to fall. No parasitaemia was observed in any of the pigs in the splenectomized and unsplenectomized uninoculated control animals. The pigs carrying either single or mixed infections had pyrexia and their temperatures were consistently higher than those of the control animals. Infected pigs also showed anaemia with significant decline in Pcv, Hb and RBC values. The total WBC count rose in the infected pigs and the rise was more pronounced in pigs carrying E. suis alone. In addition, it was only in pigs infected by E. suis alone that a decline in the percentage of neutrophils and a rise in percentage lymphocytes was observed.