Improved molecular detection of Ehrlichia and Anaplasma species applied to Amblyomma ticks collected from cattle and sheep in Ethiopia.

Detection of Ehrlichia and Anaplasma species in animals and tick vectors is crucial for an understanding of the epidemiology of diseases caused by these pathogens. In this study, a pair of primers designated EBR2 and EBR3 was designed from the Anaplasma 16S rDNA sequence and was used along with a previously described primer EHR 16SD for the simultaneous detection of Ehrlichia and Anaplasma species by nested PCR. The primers were used to amplify 925bp of DNA from known species of Ehrlichia and Anaplasma. Restriction with MboII and MspI enzymes allowed Ehrlichia and Anaplasma speciation. Restriction with MboII differentiated between An. marginale, Anaplasma (formerly Ehrlichia) sp. Omatjenne, and An. centrale with An. marginale and Anaplasma (formerly Ehrlichia) sp. Omatjenne yielding 2 distinct fragments each while An. centrale produced 3 distinct bands. Ehrlichia ruminantium and An. phagocytophylum remained undigested. Subsequent restriction with MspI differentiated E. ruminantium from An. phagocytophylum with 2 and 4 fragments, respectively. When used on tick samples from the field, 63 ticks (16.4%) out of 384 collected from cattle and sheep were positive for one or more species of Ehrlichia and Anaplasma. The positivity ranged from 6.3% at Andasa to 36.7% at Habernosa. Higher overall infection rates were found in Amblyomma lepidum than in Amblyomma variegatum ticks (p=0.009). Amblyomma lepidum from Habernosa were more often infected with all detected species of Anaplasma and Ehrlichia than Am. variegatum. At Bako, however, Anaplasma (formerly Ehrlichia) sp. Omatjenne was detected only in Am. variegatum. A significantly higher proportion of ticks collected from cattle (20.6%) was found positive than in those collected from sheep (3.3%) (p=0.003). Simultaneous detection of Ehrlichia and Anaplasma species and correct identification of mixed infections was possible. Since the ticks were collected from animals, the occurrence of the major species of Ehrlichia and Anaplasma in ruminants in the area is confirmed.

African horse sickness outbreaks caused by multiple virus types in Ethiopia.

African horse sickness (AHS) is associated with high morbidity and mortality in equids, especially horses. A retrospective analysis was carried out concerning 737 AHS outbreaks that occurred during 2007-2010 in Ethiopia. A total of ten outbreaks were investigated in the study period. All four forms of the disease (pulmonary, cardiac, horse sickness fever and the combined form) were observed, with the cardiac form being the most prevalent. Multiple African horse sickness virus serotypes (AHSV-2, AHSV-4, AHSV-6, AHSV-8 and AHSV-9) were detected by molecular methods (type-specific real-time RT-PCR assays), and fourteen isolates were derived from blood and tissue samples collected during 2009-2010. This is the first report of AHSV-4, AHSV-6 or AHSV-8 in Ethiopia.

Epidemiological study on foot-and-mouth disease in cattle: seroprevalence and risk factor assessment in South Omo zone, south-western Ethiopia.

A cross-sectional sero-epidemiological study was conducted in seven districts of the South Omo zone, south-western Ethiopia, between October 2008 and May 2009 with the objective of determining the seroprevalence of foot-and-mouth disease (FMD) in cattle and identifying the potential risk factors associated with the disease. In total, 770 cattle sera samples were collected and submitted to the National Veterinary Institute (NVI), Debre Zeit, Ethiopia, for screening using the 3ABC-ELISA. The overall seroprevalence of 8.18% (n=63) was recorded in the study. The highest district-level prevalence was observed in Bennatsemay district (30.2%), and the lowest prevalence was in Malle and Debub Aari districts, each with prevalence of 6.3%. The difference in seropositivity of FMD in the studied districts was found to be statistically significant. From the various risk factors analysed, age of animal, contact history with wild animals, distance of the herd from parks and wild animals' sanctuary and movement pattern of herds in search of pasture and water from area to area were found to be significantly associated (P<0.05) with the seroprevalence of FMD. The results of this study showed that FMD is an important cattle disease in the study areas. Thus, an appropriate control strategy has to be designed and applied, which could involve regulation of transboundary cattle movement, prevention of contact with wildlife and vaccination against the circulating virus strain.

Bovine mastitis and its association with selected risk factors in smallholder dairy farms in and around Bahir Dar, Ethiopia.

Three hundred fifty one (195 local zebu and 156 Holstein x local zebu crosses) lactating cows of smallholder farms in Bahir Dar 'milk shed' were examined from September 2003 to March 2004 to determine mastitis prevalence, isolate pathogens and identify the role of some potential risk factors. Clinical prevalence was determined through examination of abnormalities of milk, udder or cow. California mastitis test (CMT) was used for determination of subclinical mastitis prevalence. Clinical prevalence at cow level was 3.9% in crossbreds and none in local zebu breeds. Subclinical mastitis at cow level based on CMT was high (34.4%) in crossbreds compared to indigenous zebu (17.9%) (p < 0.05). Quarter subclinical prevalence based on CMT was 17.9% and 4.9% in crossbreds and local zebu, respectively. The pathogens isolated from mastitic milk (CMT positive milk) were coagulase negative staphylococci (CNS), S. aureus, Str. agalactiae, Str. dysgalactiae, Str. uberis, Micrococcus species, C. bovis, A. pyogens, B. cereus, and S. intermedius. Among these, the most frequent isolates were CNS (50%), S. aureus (19%), Str. agalactiae (8%) and Str. dysgalactiae (7%). Among potential risk factors considered, stage of lactation, parity and breed were found to affect the occurrence of mastitis significantly (p < 0.05).

Pharmacokinetics and safety of nelfinavir when used in combination with zidovudine and lamivudine in HIV-infected pregnant women: Pediatric AIDS Clinical Trials Group (PACTG) Protocol 353.

BACKGROUND: Combination antiretroviral regimens including nelfinavir (NFV) are commonly used in pregnancy. We studied the safety, antiviral effect, and pharmacokinetics of NFV and its M8 metabolite with two dosing regimens in combination with zidovudine (ZDV) and lamivudine (3TC) in HIV-infected pregnant women. METHOD: HIV-infected pregnant women between 14 and 34 weeks gestation received NFV (Cohort 1: 750 mg tid, n = 10; Cohort 2: 1250 mg bid, n = 23) with ZDV and 3TC. Serial blood sampling for NFV concentrations was performed antepartum (AP) and 6 weeks postpartum (PP). Maternal and cord blood samples were also obtained at delivery. NFV and M8 levels were determined by high-performance liquid chromatography. The pharmacokinetic (PK) target was an extrapolated NFV AUC0-24 > 30 mug . h/mL. Mothers were followed frequently for potential clinical and laboratory toxicity. RESULTS: Overall, NFV in combination with ZDV and 3TC was well tolerated. The PK target was met in 3/8 AP and 5/7 PP in Cohort 1 and 17/21 AP and 16/17 PP in Cohort 2. When Cohort 2 NFV PK parameters AP and PP were compared, median Cmax (3.90 microg/mL vs. 5.01 microg/mL, p < .05) and AUC0-24 (56.6 vs. 86.8 microg . h/mL, p < .05) were increased PP and oral clearance (Cl/F; 44.2 vs. 28.8 L/h, p < .05) was decreased PP. The average M8/NFV ratio was increased PP compared to AP (0.085 vs. 0.29, p < .001). Placental transfer of NFV was low with a median cord blood:maternal plasma ratio at delivery of 0.05. Maternal mean CD4+ T cell counts increased significantly and plasma HIV-1 RNA levels decreased from entry to delivery and 6 to 12 weeks postpartum. CONCLUSION: NFV used in combination with ZDV and 3TC was well tolerated in pregnant HIV-infected women and produced a significant improvement in HIV disease parameters. NFV drug exposure is inadequate in most pregnant women receiving 750 mg tid but is much improved with 1250 mg bid. NFV crosses the placenta poorly. The AP increase in NFV oral clearance and decrease in M8/NFV ratio suggest that CYP3A activity increases relative to CYP2C19 activity during pregnancy.

A study on contagious caprine pleuropneumonia (CCPP) in goats at an export oriented abattoir, Debrezeit, Ethiopia.

300 goat serum samples from an export-oriented abattoir were tested for contagious caprine pleuropneumonia antibodies by the complement fixation test. The disease prevalence was 31% with no significant differences (P > 0.05) between the regions "Borena", "Bale", "Afar" and "Jinka" or the age of the goats (P > 0.05). Gross pathology and histopathology of the lung primary lesions were indicative of pleuropneumonia caused by Mycoplasma capricolum subsp. capripneumoniae.

Distribution and superinfection of bovine leukemia virus genotypes in Japan.

A study to investigate the types and distribution of bovine leukemia virus (BLV) was conducted on about eight hundred cattle drawn from 53 farms found in 16 prefectures in Japan. Agar gel immunodiffusion (AGID) tests of serum samples and nested-PCR to detect BLV provirus, in peripheral blood leukocytes were performed. To identify genotypes, restriction fragment length polymorphism (RFLP) was performed with a PCR-amplified 444 bp fragment of the env gene using endonucleases. Three genotypes (1, 3, and 5) were dominant in Japan, and were found in 48.3%, 32.7%, and 16.9% of PCR positive cattle, respectively. Of the cattle infected with genotype 1, 84.7% were strongly positive in the AGID test. Similarly, in cattle with genotype 3, 78.9% were strongly positive. However, only 59.1% of cattle with genotype 5 were strong positive. Three cattle showed unusual RFLP patterns and they were found to be infected with more than one genotype. These results suggest that some BLV infected cattle can not induce effective immune reactions and suffer from superinfection by BLV in the field.