Reduction of Trypanosoma equiperdum from equine semen by single layer centrifugation.

Trypanosoma equiperdum (T. equiperdum) causes dourine, a venereally transmitted infection in horses. Purification of semen by single layer centrifugation (SLC) has been proven to be successful in reducing venereally transmitted diseases when dealing with other pathogens. The objective of this study was to evaluate the purification of T. equiperdum spiked semen by SLC. Semen was spiked using cryopreserved T. equiperdum stabilates (Dodola strain isolate 943). In total, 6 concentrations, varying from 102 to >5 × 106 trypanosomes, were added to semen samples. Subsequently, SLC was performed following standard procedures. The presence of the parasite in the purified semen was checked by wet smear examination, ITS1 PCR and in vivo inoculation in mice. Before SLC, all spiked semen samples, except the negative controls, were positive on PCR analysis. After SLC, all the pellets were found to be negative for T. equiperdum on microscopic examinations. Examination of the pellet by PCR could also not detect any parasite-DNA in the SLC-pellet of semen spiked with the lower number of parasites (102 to104 trypanosomes). However, in the SLC pellets spiked with 104 - 5 × 104 trypanosomes, only 1 out of the 4 replicates was negative for parasite DNA. All groups spiked with >5 × 104 trypanosomes were found to be positive on PCR. All mice in the positive controls exhibited parasitaemia (5/5). Mice inoculated with SLC-purified semen that was spiked with lower than 5 × 104 trypanosomes, remained free of parasitaemia, similar to the negative controls. However inoculation with SLC-pellets from samples with a higher number of trypanosomes (>5 × 104 - 5 × 106 and > 5 × 106), induced parasitaemia in 2 out of 5 and 3 out of 5 mice, respectively. This study indicates that single layer centrifugation can be used to clear T. equiperdum infected semen but that the success is dependent on the number of parasites.

Prevalence and antimicrobial susceptibility of Escherichia coli O157 in beef at butcher shops and restaurants in central Ethiopia.

BACKGROUND: Ethiopia bears the largest burden of foodborne diseases in Africa, and diarrheal diseases are the second leading causes of premature deaths. Enterohemorrhagic Escherichia coli O157 causes an asymptomatic infection to severe diarrhea and/or hemolytic-uremic syndrome in humans. METHODS: A total of 440 beef carcass and in-contact surface swabs from 55 butcher shops and 85 minced beef samples from 40 restaurants in central Ethiopia were collected and examined for the presence of E. coli O157. Standard microbiological methods were used to isolate and identify E. coli O157 and to characterize the antimicrobial resistance of the isolates. RESULTS: E. coli O157 was detected in 4.5% carcass swabs (n = 5) and 3.6% cutting board swabs (n = 4) samples from butcher shops. E. coli O157 was not detected in any of the minced beef samples obtained from restaurants. All isolates (n = 9) were 100% susceptible to five drugs, but five isolates were resistant to amoxicillin, two isolates to streptomycin and three isolates to chloramphenicol. One isolate was resistant to two drugs and another to three drugs. CONCLUSIONS: The present study shows a low prevalence of E. coli O157 in beef sold at butcher shops. Nevertheless, given the low infective dose of this pathogen and the deep-rooted tradition of consuming raw or undercooked beef, the current prevalence should not be considered lightly from a public health perspective.

Prevalence of Escherichia coli O157:H7 in beef cattle at slaughter and beef carcasses at retail shops in Ethiopia.

BACKGROUND: There is paucity of information regarding the epidemiology of Escherichia coli O157: H7 in developing countries. In this study, we investigated the occurrence of E. coli O157: H7 associated with beef cattle at processing plants and at retail shops in Ethiopia. METHODS: Various samples were collected from beef cattle at slaughter/processing plants, carcass at retail shops and humans at health centers. E. coli O157: H7 was isolated, identified and characterized for antimicrobial resistance, using standard microbiological methods. RESULTS: At the processing plants E. coli O157: H7 was detected in 1.89% of fecal, 0.81% of intestinal mucosal swab, 0.54% of skin swab and 0.54% of carcass internal swab samples. At retail shops it was detected in 0.8% of carcass and 0.8% of cutting board swab samples, while all samples from utensils, hands from workers, and fecal and stool samples were negative. All isolates were resistant to Amoxicillin, moderately resistant to Cefoxitine and Nitrofurantoins but susceptible to other antimicrobials tested. CONCLUSIONS: E. coli O157: H7 occurs at low prevalence in beef cattle, and the current sanitary dressing procedures in the processing plants and storage conditions in the retail shops are effective against E. coli O157: H7.

Orally fed seeds producing designer IgAs protect weaned piglets against enterotoxigenic Escherichia coli infection.

Oral feed-based passive immunization can be a promising strategy to prolong maternal lactogenic immunity against postweaning infections. Enterotoxigenic Escherichia coli (ETEC)-caused postweaning diarrhea in piglets is one such infection that may be prevented by oral passive immunization and might avert recurrent economic losses to the pig farming industry. As a proof of principle, we designed anti-ETEC antibodies by fusing variable domains of llama heavy chain-only antibodies (VHHs) against ETEC to the Fc part of a porcine immunoglobulin (IgG or IgA) and expressed them in Arabidopsis thaliana seeds. In this way, four VHH-IgG and four VHH-IgA antibodies were produced to levels of about 3% and 0.2% of seed weight, respectively. Cotransformation of VHH-IgA with the porcine joining chain and secretory component led to the production of light-chain devoid, assembled multivalent dimeric, and secretory IgA-like antibodies. In vitro analysis of all of the antibody-producing seed extracts showed inhibition of bacterial binding to porcine gut villous enterocytes. However, in the piglet feed-challenge experiment, only the piglets receiving feed containing the VHH-IgA-based antibodies (dose 20 mg/d per pig) were protected. Piglets receiving the VHH-IgA-based antibodies in the feed showed a progressive decline in shedding of bacteria, significantly lower immune responses corroborating reduced exposure to the ETEC pathogen, and a significantly higher weight gain compared with the piglets receiving VHH-IgG producing (dose 80 mg/d per pig) or wild-type seeds. These results stress the importance of the antibody format in oral passive immunization and encourage future expression of these antibodies in crop seeds.

Establishing mono-eukaryotic Histomonas meleagridis cultures from in vivo infection contaminated with Tetratrichomonas gallinarum and Blastocystis spp.

SUMMARY The necessity to easily establish Histomonas meleagridis cultures has been underlined extensively by many researchers in order to gain more insights in the biology of H. meleagridis. In addition the occurrence of different protozoa in the caeca of birds impedes, however, the isolation and propagation of H. meleagridis from field outbreaks. Therefore, in a kinetic study using transmission electron microscopy the deleterious effects of adventitious protozoa including Tetratrichomonas gallinarum and Blastocystis spp. on cultured H. meleagridis were examined. To overcome this issue, an easy and successful approach to establish the mono-eukaryotic H. meleagridis culture free of other host's protozoa is proposed. At 10 days post infection, liver lesions of H. meleagridis-infected birds were isolated and inoculated into culture media pre-incubated with caecal bacteria. After 48 h of incubation, presence of H. meleagridis in the cultures was confirmed through morphological evaluation. Additionally, TEM examination and analysis by PCR amplification of the small subunit rRNA gene could exclude the co-cultivation of T. gallinarum and Blastocystis spp. Furthermore, after successful propagation and maintenance of the cultured H. meleagridis, its pathogenicity was affirmed in an infection experiment in turkeys.

Several enteropathogens are circulating in suckling and newly weaned piglets suffering from diarrhea in the province of Villa Clara, Cuba.

Intestinal contents of suckling (n = 45) and newly weaned (n = 45) piglets, suffering from diarrhea in the province of Villa Clara in Cuba, were tested for viral, bacterial, and parasitic enteropathogens from May to June 2008. At least one enteropathogen was identified in 53.3 % of piglets and enterotoxigenic Escherichia coli (ETEC; 25.6 %) was the major pathogen; mostly STa(+)/STb(+) or F4(+)/STa(+)/STb(+) ETEC were isolated. The overall occurrence of the rest of pathogens was 10 % for transmissible gastroenteritis virus (TGEV) and Cryptosporidium parvum, 6.7 % for rotavirus A and Isospora suis, 5.6 % for α-toxigenic Clostridium perfringens, 3.3 % for verotoxigenic E. coli (VTEC), and 2.2 % for Salmonella enterica subspecies enterica serovar Newport. TGEV and α-toxigenic C. perfringens were only identified in suckling piglets, while Salmonella Newport and VTEC were only detected in weaned pigs. Porcine epidemic diarrhea virus (PEDV), ß-toxigenic C. perfringens, Eimeria spp., and helminths were not identified. Eight kinds of mixed infections were detected in 25 % of enteropathogen positive piglets. ETEC was present in 10 of 12 mixed infections, and TGEV infections were never combined. This survey demonstrates that several enteropathogens are circulating in piggeries located in the province of Villa Clara in Cuba, and that is necessary to improve surveillance, prevention, and control of enteric infections in order to increase production efficiency.

Protection of piglets against Edema disease by maternal immunization with Stx2e toxoid.

Edema disease (ED) in piglets is caused by Shiga toxin Stx2e-producing Escherichia coli. We show that a genetically disarmed Stx2e toxoid is a safe antigen that generates antiserum protecting piglets against the Stx2e toxin. Immunization of suckling piglets with the Stx2e toxoid was safe, had no adverse effects on growth of the piglets, and resulted in effective prevention of edema disease clinical symptoms after challenge with the Stx2e toxin. Our data showed that maternal immunity against the Stx2e toxoid can be transmitted from the vaccinated sows to the piglets via the colostrum. Very high levels of Stx2e-specific serum antibodies persisted in these piglets until 1 month postweaning, bridging the critical period in which the weaned piglets are most susceptible to edema infection. Challenge with Stx2e toxin resulted in clinical signs of edema disease and death of all control piglets from nonimmunized sows, whereas none of the piglets from immunized sows developed clinical signs of ED.

Identification of Avian pathogenic Escherichia coli genes that are induced in vivo during infection in chickens.

Avian pathogenic Escherichia coli (APEC) is associated with extraintestinal infections in poultry causing a variety of diseases collectively known as colibacillosis. The host and bacterial factors influencing and/or responsible for carriage and systemic translocation of APEC inside the host are poorly understood. Identification of such factors could help in the understanding of its pathogenesis and in the subsequent development of control strategies. Recombination-based in vivo expression technology (RIVET) was used to identify APEC genes specifically expressed during infection in chickens. A total of 21 clones with in vivo-induced promoters were isolated from chicken livers and spleens, indicative of systemic infection. DNA sequencing of the cloned fragments revealed that 12 of the genes were conserved E. coli genes (metH, lysA, pntA, purL, serS, ybjE, ycdK [rutC], wcaJ, gspL, sdsR, ylbE, and yjiY), 6 of the genes were phage related/associated, and 3 genes were pathogen specific (tkt1, irp2, and eitD). These genes are involved in various cellular functions, such as metabolism, cell envelope and integrity, transport systems, and virulence. Others were phage related or have yet-unknown functions.

High prevalence of F4+ and F18+ Escherichia coli in Cuban piggeries as determined by serological survey.

Little information is available on the prevalence of swine enteropathogens in Cuba where diarrheic diseases are responsible for 31% and 37% of the total mortality during the neonatal and postweaning periods. F4+ and F18+ enterotoxigenic Escherichia coli and F18+ verotoxigenic E. coli induce diarrhea and edematous disease in pigs, but their distribution has never been thoroughly studied in the Cuban swine population. Therefore, the present study estimated the prevalence of F4- and F18-specific antibodies in sera of 1,044 6-month-old gilts distributed in 34 piggeries spread over the Cuban territory. For the data analysis, which included the optical density of individual samples tested by ELISA, random-effects models and a mixture model in R (package "mixAK"; Komárek, Computational Statistics and Data Analysis 53:3932-3947, 2009) were fitted. Low, moderate, and high levels of F4-specific antibodies were found in 67.6%, 26.8%, and 5.6% of the gilts, while 66.4% and 33.6% of gilts showed low and high levels of F18-specific antibodies. Hereby, we show that F4+ and F18+ E. coli are highly prevalent as potential enteropathogens in Cuban piggeries.

Escherichia coli strains causing edema disease in northern Vietnam share an identical verotoxin 2e.

Edema disease (ED) is a common fatal disease in newly weaned piglets. To develop an effective control program for ED, we carried out a study to better understand the incidence and spread of the disease and the characteristics of the causative agent. In our study, 69 Escherichia coli strains, isolated from 92 piglets showing clinical signs of ED from 13 provinces in northern Vietnam, were positive for both the VT2e toxin and the F18 major fimbrial subunit gene fedA. Of these, 40 strains (58%) were positive for AIDA and 16 isolates carried one or more enterotoxins. Forty-six (67%) of the 69 VT2e(+)/F18(+) E. coli isolates belonged to classical serotypes (O139:K82, O141: K85, O138:K81, and O149:K91) while the remaining strains did not belong to the common serotypes in pig. Seropathotype 0139:K82(+)/VT2e(+)/F18(+)/AIDA(+) (21 isolates) was the most frequently detected ED-causing E. coli strain. High prevalence of resistance was observed to the common drugs of tetracycline, streptomycin, trimethoprim/sulfamethoxazole, amoxicillin/clavulanic acid, and spectinomycin. Multiple resistances were widely distributed with 84% of isolates resistant to five antibiotics. Sequence analysis demonstrated that the VT2e toxin is identical among E. coli strains causing ED in pig.

Evaluation of the persistence and gene expression of an anti-Chlamydophila psittaci DNA vaccine in turkey muscle.

BACKGROUND: DNA vaccination has been shown to elicit specific cellular and humoral immune responses to many different agents in a broad variety of species. However, looking at a commercial use, the duration of the immune response against the vaccine is critical. Therefore the persistence of the DNA vaccine, as well as its expression, should be investigated. We conducted these investigations on a DNA vaccine against Chlamydophila psittaci, a Gram-negative intracellular bacterium which causes respiratory disease in turkeys and humans. Previous studies showed that the DNA vaccine confers partial protection against C. psittaci infection in turkeys. Turkeys were injected intramuscularly with the DNA vaccine : a eukaryotic expression vector (pcDNA1::MOMP) expressing the major outer membrane protein (MOMP) of an avian C. psittaci serovar D strain. Over a period of 11 weeks, cellular uptake of the DNA vaccine was examined by PCR, transcription of the insert by reverse transcript-PCR (RT-PCR) and mRNA translation by immunofluorescence staining of muscle biopsies. RESULTS: The results indicate that the DNA vaccine persists in turkey muscle for at least 10 weeks. Moreover, during this period of time MOMP was continuously expressed, as evidenced by the immunofluorescence staining and RT-PCR. CONCLUSION: Since C. psittaci infections occur at the age of 3 to 6 and 8 to 12 weeks, a vaccine persistence of 10 weeks seems adequate. Therefore, further research should concentrate on improving the elicited immune response, more specifically the cell-mediated immune response, rather than prolonging the lifespan of the plasmid.

New Trypanosoma evansi Type B Isolates from Ethiopian Dromedary Camels.

BACKGROUND: Trypanosoma (T.) evansi is a dyskinetoplastic variant of T. brucei that has gained the ability to be transmitted by all sorts of biting flies. T. evansi can be divided into type A, which is the most abundant and found in Africa, Asia and Latin America and type B, which has so far been isolated only from Kenyan dromedary camels. This study aimed at the isolation and the genetic and phenotypic characterisation of type A and B T. evansi stocks from camels in Northern Ethiopia. METHODOLOGY/PRINCIPAL FINDINGS: T. evansi was isolated in mice by inoculation with the cryopreserved buffy coat of parasitologically confirmed animals. Fourteen stocks were thus isolated and subject to genotyping with PCRs targeting type-specific variant surface glycoprotein genes, mitochondrial minicircles and maxicircles, minisatellite markers and the F1-ATP synthase γ subunit gene. Nine stocks corresponded to type A, two stocks were type B and three stocks represented mixed infections between A and B, but not hybrids. One T. evansi type A stock was completely akinetoplastic. Five stocks were adapted to in vitro culture and subjected to a drug sensitivity assay with melarsomine dihydrochloride, diminazene diaceturate, isometamidium chloride and suramin. In vitro adaptation induced some loss of kinetoplasts within 60 days. No correlation between drug sensitivity and absence of the kinetoplast was observed. Sequencing the full coding sequence of the F1-ATP synthase γ subunit revealed new type-specific single nucleotide polymorphisms and deletions. CONCLUSIONS/SIGNIFICANCE: This study addresses some limitations of current molecular markers for T. evansi genotyping. Polymorphism within the F1-ATP synthase γ subunit gene may provide new markers to identify the T. evansi type that do not rely on variant surface glycoprotein genes or kinetoplast DNA.

Maternal colostral leukocytes appear to enhance cell-mediated recall response, but inhibit humoral recall response in prime-boost vaccinated calves.

Whether colostral leukocytes (CLs) of vaccinated dams influence the immune response of neonatal calves following vaccination against the same antigen as their respective dams remains unanswered. Therefore, we compared the induction of humoral and cellular immune responses after vaccination in calves that had received CL-free or maternal CL-enriched colostrum from a cell-free colostrum bank of nonvaccinated cows. Also, vaccinated calves that had received fresh maternal colostrum from their own dam were included in the study. Moreover, we analyzed whether the post-partum time of priming vaccination (day 2, 5 or 10) of the calves could influence the outcome of the immune responses. All calves received a booster vaccination 23 days after the priming vaccination. All calves showed only an increase in tetanus toxoid (TT)-specific antibodies and TT-induced proliferation after booster vaccination. Tetanus toxoid-specific antibody responses in calves increased immediately after booster vaccination, irrespective of whether or not their cell-free bank colostrum had been enriched with CLs from their own dam. Conversely, calves receiving their own plain dam colostrum displayed a later humoral response, due to colostral antibodies. After booster vaccination, calves of the CL-enriched colostrum group had a more pronounced antigen-specific proliferative response than the calves of the CL-free colostrum group. We propose that CLs might have a suppressive influence on the emergence of the TT-specific antibodies, but an enhancing effect on the TT-specific lymphocyte proliferation of newborn calves upon TT vaccination, which is dependent on the time point of the priming vaccination.

Ribosomal DNA analysis of tsetse and non-tsetse transmitted Ethiopian Trypanosoma vivax strains in view of improved molecular diagnosis.

Animal trypanosomosis caused by Trypanosoma vivax (T. vivax) is a devastating disease causing serious economic losses. Most molecular diagnostics for T. vivax infection target the ribosomal DNA locus (rDNA) but are challenged by the heterogeneity among T. vivax strains. In this study, we investigated the rDNA heterogeneity of Ethiopian T. vivax strains in relation to their presence in tsetse-infested and tsetse-free areas and its effect on molecular diagnosis. We sequenced the rDNA loci of six Ethiopian (three from tsetse-infested and three from tsetse-free areas) and one Nigerian T. vivax strain. We analysed the obtained sequences in silico for primer-mismatches of some commonly used diagnostic PCR assays and for GC content. With these data, we selected some rDNA diagnostic PCR assays for evaluation of their diagnostic accuracy. Furthermore we constructed two phylogenetic networks based on sequences within the smaller subunit (SSU) of 18S and within the 5.8S and internal transcribed spacer 2 (ITS2) to assess the relatedness of Ethiopian T. vivax strains to strains from other African countries and from South America. In silico analysis of the rDNA sequence showed important mismatches of some published diagnostic PCR primers and high GC content of T. vivax rDNA. The evaluation of selected diagnostic PCR assays with specimens from cattle under natural T. vivax challenge showed that this high GC content interferes with the diagnostic accuracy of PCR, especially in cases of mixed infections with T. congolense. Adding betain to the PCR reaction mixture can enhance the amplification of T. vivax rDNA but decreases the sensitivity for T. congolense and Trypanozoon. The networks illustrated that Ethiopian T. vivax strains are considerably heterogeneous and two strains (one from tsetse-infested and one from tsetse-free area) are more related to the West African and South American strains than to the East African strains. The rDNA locus sequence of six Ethiopian T. vivax strains showed important differences and higher GC content compared to other animal trypanosomes but could not be related to their origin from tsetse-infested or tsetse-free area. The high GC content of T. vivax DNA renders accurate diagnosis of all pathogenic animal trypanosomes with one single PCR problematic.

The Enrichment of Histomonas meleagridis and Its Pathogen-Specific Protein Analysis: A First Step to Shed Light on Its Virulence.

Since the discovery of Histomonas meleagridis in 1893, the necessity of isolating pure H. meleagridis has been highlighted over the years in the battle against histomonosis. Insights into the molecular characteristics of this protozoon open possibilities to proper treatment. Axenization of H. meleagridis in vitro cultures cocultured with bacteria has been unsuccessful. Numerous unsuccessful attempts at culturing H. meleagridis axenically have reinforced the assumption that the protozoa had an obligate relationship with certain bacteria originating from the host ceca. Within these perspectives, we enriched H. meleagridis cells from a mono-eukaryotic culture copropagated with host cecal bacteria by flow cytometry. The enrichment of histomonads was confirmed through transmission electron microscopy and two-dimensional gel electrophoresis. For the first time several protein spots were successfully identified. The majority of spots were annotated as cytoskeletal proteins. Actin microfilaments are known to be a key player in cell spreading, cell adhesion, phagocytosis, signal transduction, and several other processes. Together with the identification of superoxide dismutase, the information generated from protein analysis of H. meleagridis may serve as a very first step toward understanding its pathogenesis and virulence.

Trypanosoma equiperdum: master of disguise or historical mistake?

After 100 years of research, only a small number of laboratory strains of Trypanosoma equiperdum exists, and the history of most of the strains is unknown. No definitive diagnosis of dourine can be made at the serological or molecular level. Only clinical signs are pathognomonic and international screening relies on an outdated cross-reactive serological test (the complement-fixation test) from 1915, resulting in serious consequences at the practical level. Despite many characterization attempts, no clear picture has emerged of the position of T. equiperdum within the Trypanozoon group. In this article, we highlight the controversies that exist regarding T. equiperdum, and the overlap that occurs with Trypanosoma evansi and Trypanosoma brucei brucei. By revisiting the published data, from the early decades of discovery to the recent serological- and molecular-characterization studies, a new hypothesis arises in which T. equiperdum no longer exists as a separate species and in which current strains can be divided into T. evansi (the historical mistake) and Trypanosoma brucei equiperdum (the master of disguise). Hence, dourine is a disease caused by specific host immune responses to a T. b. equiperdum or T. evansi infection.

Surra Sero K-SeT, a new immunochromatographic test for serodiagnosis of Trypanosoma evansi infection in domestic animals.

Trypanosoma evansi, the causative agent of surra, infects different domestic and wild animals and has a wide geographical distribution. It is mechanically transmitted mainly by haematophagous flies. Parasitological techniques are commonly used for the diagnosis of surra but have limited sensitivity. Therefore, serodiagnosis based on the detection of T. evansi specific antibodies is recommended by the World Organisation for Animal Health (OIE). Recently, we developed a new antibody detection test for the serodiagnosis of T. evansi infection, the Surra Sero K-SeT. Surra Sero K-SeT is an immunochromatographic test (ICT) that makes use of recombinant variant surface glycoprotein rVSG RoTat 1.2, produced in the yeast Pichia pastoris. In this study, we compared the diagnostic accuracy of the Surra Sero K-SeT and the Card Agglutination Test for T. evansi Trypanosomososis (CATT/T. evansi) with immune trypanolysis (TL) as reference test on a total of 806 sera from camels, water buffaloes, horses, bovines, sheep, dogs and alpacas. Test agreement was highest between Surra Sero K-SeT and TL (κ=0.91, 95% CI 0.841-0.979) and somewhat lower between CATT/T. evansi and TL (κ=0.85, 95% CI 0.785-0.922) and Surra Sero K-SeT and CATT/T. evansi (κ=0.81, 95% CI 0.742-0.878). The Surra Sero K-SeT displayed a somewhat lower overall specificity than CATT/T. evansi (94.8% versus 98.3%, χ(2)=13.37, p<0.001) but a considerably higher sensitivity (98.1% versus 84.4%, χ(2)=33.39, p<0.001). We conclude that the Surra Sero K-SeT may become an alternative for the CATT/T. evansi for sensitive detection of antibodies against T. evansi in domestic animals.

Epidemiology of Trypanosoma evansi and Trypanosoma vivax in domestic animals from selected districts of Tigray and Afar regions, Northern Ethiopia.

BACKGROUND: African animal trypanosomosis, transmitted cyclically by tsetse flies or mechanically by other biting flies, causes serious inflictions to livestock health. This study investigates the extent of non-tsetse transmitted animal trypanosomosis (NTTAT) by Trypanosoma (T.) evansi and T. vivax in domestic animals in the tsetse-free regions of Northern Ethiopia, Afar and Tigray. METHODS: A cross sectional study was conducted on 754 dromedary camels, 493 cattle, 264 goats, 181 sheep, 84 donkeys, 25 horses and 10 mules. The microhaematocrit centrifugation technique was used as parasitological test. Plasma was collected for serodiagnosis with CATT/T.evansi and RoTat 1.2 immune trypanolysis (ITL) while buffy coat specimens were collected for molecular diagnosis with T. evansi type A specific RoTat 1.2 PCR, T. evansi type B specific EVAB PCR and T. vivax specific TvPRAC PCR. RESULTS: The parasitological prevalence was 4.7% in Tigray and 2.7% in Afar and significantly higher (z = 2.53, p = 0.011) in cattle (7.3%) than in the other hosts. Seroprevalence in CATT/T.evansi was 24.6% in Tigray and 13.9% in Afar and was significantly higher (z = 9.39, p < 0.001) in cattle (37.3%) than in the other hosts. On the other hand, seroprevalence assessed by ITL was only 1.9% suggesting cross reaction of CATT/T.evansi with T. vivax or other trypanosome infections. Molecular prevalence of T. evansi type A was 8.0% in Tigray and in Afar and varied from 28.0% in horses to 2.2% in sheep. It was also significantly higher (p < 0.001) in camel (11.7%) than in cattle (6.1%), donkey (6%), goat (3.8%), and sheep (2.2%). Four camels were positive for T. evansi type B. Molecular prevalence of T. vivax was 3.0% and was similar in Tigray and Afar. It didn't differ significantly among the host species except that it was not detected in horses and mules. CONCLUSIONS: NTTAT caused by T. vivax and T. evansi, is an important threat to animal health in Tigray and Afar. For the first time, we confirm the presence of T. evansi type B in Ethiopian camels. Unexplained results obtained with the current diagnostic tests in bovines warrant particular efforts to isolate and characterise trypanosome strains that circulate in Northern Ethiopia.

Trypanosome infection in dromedary camels in Eastern Ethiopia: Prevalence, relative performance of diagnostic tools and host related risk factors.

A cross-sectional study was conducted in Chifra and Dewe districts of Afar region, Eastern Ethiopia, to determine the prevalence, agreement between diagnostic tests and host related risk factors of trypanosome infection in camel. An overall prevalence of 2%, 24.1%, 21.3%, 9.5% and 7.8% was recorded with respectively Giemsa stained thin blood smear, CATT/T. evansi, RoTat1.2 PCR, 18S PCR and ITS-1PCR in a cohort of 399 animals. Only one T. vivax infection was confirmed by TvPRAC PCR indicating T. evansi as the predominant species affecting camels in the study area. No single animal was positive when tested with T. evansi type B specific EVAB PCR. There was slight agreement between the CATT/T. evansi and the molecular tests. Among the PCR methods, RoTat 1.2 PCR yielded a significantly higher positivity rate compared to 18S PCR and ITS-1 PCR. There was no significant difference in the positivity rate observed in each gender of camels (p>0.05). The positivity rate was significantly higher in camels with poor body condition and in older animals when tested using the CATT/T.evansi or RoTat 1.2 PCR (p>0.05). Camels that tested positive with all tests had significantly lower PCV's (p<0.05). This study provides further evidence that T. evansi is endemic in the Afar region of Ethiopia. The latent class analysis indicated an estimate overall prevalence of 19% (95% CI: 13-28). Moreover, the model indicated low sensitivity of CATT/T. evansi (43%) and the PCR tests (39-53%) but higher specificity of the PCR tests (86-99%) and low specificity of CATT/T. evansi (80%). This study suggests that improved sensitivity and reliability of the tests would help diagnosis of trypanosomosis. Further studies are required to determine the prevalence of clinical disease and losses due to trypanosomosis.

Predominance of the papGII allele with high sequence homology to that of human isolates among avian pathogenic Escherichia coli (APEC).

Avian pathogenic Escherichia coli (APEC) are often found in poultry and are responsible for a set of diseases, commonly referred to as avian colibacillosis. One of the important virulence factors is adhesion to different epithelial surfaces, which is mediated by pili. P pili are thought to play a role by means of their PapG adhesin, which occurs in three molecular variants: PapGI, PapGII and PapGIII. This study is the first to determine and analyse the distribution of the different papG alleles in APEC. Our results show a significant predominance of the papGII allele above all other alleles or allele combinations. No statistically significant associations could be found between papG allele distribution and the type of bird, organ of isolation and O serogroup. Finally, the papGII and papGIII sequences showed high homology with mammalian (including human) source papG sequences.