Purification and identification of a new antifungal dipeptide from Bacillus velezensis AR1 culture supernatant.

BACKGROUND: Bacillus velezensis and other Bacillus species are known for the production of bioactive secondary metabolites that are active against plant pathogenic fungi. In view of this, a study was undertaken to investigate the antifungal potential of B. velezensis AR1 culture supernatant. RESULT: The butanol extract concentrate of B. velezensis AR1 was separated into different fractions by column chromatography. A fraction eluted by 9:1 chloroform: methanol caused 25.8-70.2% and 25.2-56.3% growth inhibition of Monilinia fructicola and Colletotricum goeosporioides, respectively. This fraction was subjected to solid-phase extraction using a Strata SI-1 column and further purified by prep-TLC to obtain a pure metabolite showing a single peak on high performance liquid chromatography. On the basis of the nuclear magnetic resonance (NMR 1 H, 13 C, HH COSY, HSQC and HMBC) analysis, the metabolite was identified as 5-N-tyrosinylornithine. The compound exhibited antifungal activity against two plant pathogenic fungi. CONCLUSION: 5-N-tyrosinylornithine, the secondary metabolite isolated from the culture supernatant of B. velezensis AR1 exhibited significant antifungal activity against two plant pathogenic fungi.

Nutritional and metabolic implications of replacing cornstarch with D-xylose in broiler chickens fed corn and soybean meal-based diet.

Effects of substituting cornstarch with D-xylose on growth performance, nutrients digestibility, serum metabolites, and expression of select hepatic genes involved in glucose and lipid metabolism were investigated in broiler chickens. A total of 360 one-day-old male Ross chicks were fed 3 diets (n = 24; 5 chicks/cage) for 21 days. A control corn-soybean meal-based diet with 25% cornstarch was formulated to meet specifications. Two additional diets were formulated by substituting cornstarch with 5 or 15% D-xylose w/w. Growth performance and digestibility by index method were determined in 12 replicate cages. Birds in these replicates had free access to feed and water, the BW and feed intake (FI) were monitored weekly and the excreta samples were collected on d 18 to 20. The other 12 replicates were used for blood and liver sampling by serial slaughter. On d 18, baseline (t0) birds were sampled following a 12 h overnight fasting and birds allowed 30 min access to the feed; samples were subsequently taken at 60, 120, 180, 240, and 300 min post feeding. Serum metabolites (glucose, xylose, and insulin) were assayed at all time points, whereas expression of hepatic transcripts was evaluated at zero, 180 and 300 min. Xylose linearly reduced (P < 0.05) FI, BWG, gross energy digestibility, and feed conversion ratio (FCR) but increased (P < 0.05) serum xylose level. Serum glucose and insulin levels were higher (P < 0.05) in the post-fed state compared with baseline, irrespective of treatments. There was an interaction (P < 0.05) between diet and sampling time on the expression of hepatic genes. At t0, xylose linearly increased (P < 0.05) the expression of pyruvate carboxylase, Acetyl Co-A acethyltransferase 2 (ACAT2), and glucose transporter 2. Xylose linearly reduced (P < 0.05) the expression of ACAT2 at 300 min post feeding. In conclusion, 5% or more xylose reduced growth performance and utilization of nutrients linked to hepatic enzymes and transcription factors involved in glucose and lipid metabolism.

Effect of supplementing a fibrous diet with a xylanase and ß-glucanase blend on growth performance, intestinal glucose uptake, and transport-associated gene expression in growing pigs.

The present study evaluated supplemental carbohydrase effect on performance, intestinal nutrient uptake, and transporter mRNA expressions in growing pigs offered a high-fiber diet manufactured with distillers dried grains with solubles (DDGS). Twenty-four pigs (22.4 ± 0.7 kg BW) were randomly assigned to 1of 3 nutritionally adequate diets (8 pigs per diet) based on corn and soybean meal (SBM) with either 0 (control) or 30% DDGS (high fiber [HF]). The third diet was supplemented with a xylanase and ß-glucanase blend (XB) in addition to the 30% DDGS (HF+XB). Parameters determined were ADFI, ADG, G:F, plasma glucose and plasma urea nitrogen (PUN) concentrations, jejunal tissue electrophysiological properties, and mRNA expressions of the sodium-dependent glucose transport 1 (SGLT1) and cationic AA transporter, bo,+AT, in the jejunal and ileal tissues. In addition, mRNA expressions of the short-chain fatty acid transporters, monocarboxylate transporter 1 (MCT1) and sodium-coupled monocarboxylate transporter, and mucin genes were quantified in the ileum. Feed intake, plasma glucose, and jejunal tissue electrophysiological properties were not affected (P > 0.05) by diet. However, control-fed pigs had superior growth rate and feed efficiency and higher PUN (P < 0.05) than HF- and HF+XB-fed pigs. The HF diet increased (P < 0.05) SGLT1 mRNA expression in the jejunum and decreased (P < 0.05) bo,+ mRNA expression in the ileum. The XB supplementation also increased bo,+ mRNA expression in the ileum relative to HF-fed pigs. Additionally, MCT1 mRNA expression was greater (P < 0.05) in the ileum of the HF- and HF+XB-fed pigs. In the present study, XB supplementation influenced nutrient transporter mRNA expression, although it was not accompanied by improved pig performance.

Seroepidemiological study of livestock brucellosis in a pastoral region.

A seroepidemiological study of Brucella infections in multiple livestock species in the Borana pastoral system of Ethiopia was performed between December 2007 and October 2008. A cross-sectional multi-stage sampling technique was employed to select 575 cattle, 1073 camels and 1248 goats from the target populations. Sera were collected from the animals, and serially tested using Rose Bengal test and complement fixation test. Overall prevalence and prevalence with respect to explanatory variables were established, and potential risk factors for seropositivity were analysed using a multivariable logistic regression. The results showed that 8·0% (95% CI 6·0-10·6), 1·8% (95% CI 1·1-2·8) and 1·6% (95% CI 1·0-2·5) of the tested cattle, camels and goats, respectively, had antibodies to Brucella antigen. Positive reactors were found in 93·8% of the villages with more frequent detection of positive cattle (93·3%) than camels (56·3%) and goats (37·5%). Risk factors identified for cattle were: keeping more livestock species at household level (OR 4·1, 95% CI 1·9-8·9), increasing age of the animal (OR 2·8, 95% CI 1·3-6·0) and wet season (OR 3·3, 95% CI 1·6-6·9). Increase in household-level species composition (OR 4·1, 95% CI 1·2-14·2) and wet season (OR 3·7, 95% CI 1·5-9·1) were found to be risk factors for seropositivity in camels and goats, respectively. Existence of more than one seroreactor animal species in most villages and association of increased livestock species composition with seropositivity may add more credence to the possibility of cross-species transmission of Brucella infections. Although no attempt to isolate Brucella spp. was made, our results suggest that cattle are more likely maintenance hosts of Brucella abortus which has spread to goats and camels. This should be substantiated by further isolation and identification of Brucella organisms to trace the source of infection and transmission dynamics in various hosts kept under mixed conditions. In conclusion, the present study suggests the need for investigating a feasible control intervention and raising public awareness on prevention methods of human exposure to brucellosis.

Transcriptome dynamics and molecular cross-talk between bovine oocyte and its companion cumulus cells.

BACKGROUND: The bi-directional communication between the oocyte and its companion cumulus cells (CCs) is crucial for development and functions of both cell types. Transcripts that are exclusively expressed either in oocytes or CCs and molecular mechanisms affected due to removal of the communication axis between the two cell types is not investigated at a larger scale. The main objectives of this study were: 1. To identify transcripts exclusively expressed either in oocyte or CCs and 2. To identify those which are differentially expressed when the oocyte is cultured with or without its companion CCs and vice versa. RESULTS: We analyzed transcriptome profile of different oocyte and CC samples using Affymetrix GeneChip Bovine Genome array containing 23000 transcripts. Out of 13162 genes detected in germinal vesicle (GV) oocytes and their companion CCs, 1516 and 2727 are exclusively expressed in oocytes and CCs, respectively, while 8919 are expressed in both. Similarly, of 13602 genes detected in metaphase II (MII) oocytes and CCs, 1423 and 3100 are exclusively expressed in oocytes and CCs, respectively, while 9079 are expressed in both. A total of 265 transcripts are differentially expressed between oocytes cultured with (OO+CCs) and without (OO-CCs) CCs, of which 217 and 48 are over expressed in the former and the later groups, respectively. Similarly, 566 transcripts are differentially expressed when CCs mature with (CCs+OO) or without (CCs-OO) their enclosed oocytes. Of these, 320 and 246 are over expressed in CCs+OO and CCs-OO, respectively.While oocyte specific transcripts include those involved in transcription (IRF6, POU5F1, MYF5, MED18), translation (EIF2AK1, EIF4ENIF1) and CCs specific ones include those involved in carbohydrate metabolism (HYAL1, PFKL, PYGL, MPI), protein metabolic processes (IHH, APOA1, PLOD1), steroid biosynthetic process (APOA1, CYP11A1, HSD3B1, HSD3B7). Similarly, while transcripts over expressed in OO+CCs are involved in carbohydrate metabolism (ACO1, 2), molecular transport (GAPDH, GFPT1) and nucleic acid metabolism (CBS, NOS2), those over expressed in CCs+ OO are involved in cellular growth and proliferation (FOS, GADD45A), cell cycle (HAS2, VEGFA), cellular development (AMD1, AURKA, DPP4) and gene expression (FOSB, TGFB2). CONCLUSION: In conclusion, this study has generated large scale gene expression data from different oocyte and CCs samples that would provide insights into gene functions and interactions within and across different pathways that are involved in the maturation of bovine oocytes. Moreover, the presence or absence of oocyte and CC factors during bovine oocyte maturation can have a profound effect on transcript abundance of each cell types, thereby showing the prevailing molecular cross-talk between oocytes and their corresponding CCs.

Suppression of the transcription factor MSX1 gene delays bovine preimplantation embryo development in vitro.

This study was conducted to investigate the effect of suppressing transcription factor gene MSX1 on the development of in vitro produced bovine oocytes and embryos, and identify its potential target genes regulated by this gene. Injection of long double-stranded RNA (LdsRNA) and small interfering RNA (siRNA) at germinal vesicle stage oocyte reduced MSX1 mRNA expression by 73 and 37% respectively at metaphase II stage compared with non-injected controls. Similarly, injection of the same anti-sense oligomers at zygote stage reduced MSX1 mRNA expression by 52 and 33% at 8-cell stage compared with non-injected controls. Protein expression was also reduced in LdsRNA- and siRNA-injected groups compared with non-injected controls at both stages. Blastocysts rates were 33, 28, 20 and 18% in non-injected control, scrambled RNA (scRNA), LdsRNA- and siRNA-injected groups respectively. Cleavage rates were also significantly reduced in Smartpool siRNA (SpsiRNA)-injected group (53.76%) compared with scRNA-injected group (57.76%) and non-injected control group (61%). Large-scale gene expression analysis showed that 135 genes were differentially regulated in SpsiRNA-injected group compared with non-injected controls, of which 54 and 81 were down- and up-regulated respectively due to suppression of MSX1. Additionally, sequence homology mapping and gene enrichment analysis with known human pathway information identified several functional modules that were affected due to suppression of MSX1. In conclusion, suppression of MSX1 affects oocyte maturation, embryo cleavage rate and the expression of several genes, suggesting its potential role in the development of bovine preimplantation embryos.

Major metacestodes in cattle slaughtered at Wolaita Soddo Municipal abattoir, Southern Ethiopia: prevalence, cyst viability, organ distribution and socioeconomic implications.

A cross sectional study to determine the prevalence and socioeconomic importance of major metacestodes of cattle was conducted from November 2007 to April 2008 at Wolaita Soddo abattoir. Accordingly, of 415 randomly selected slaughtered cattle, 11.3% and 15.4% were infected with Cysticercus bovis and hydatid cysts, respectively. The anatomical distributions of these parasites include various organs. The major risk factors for cysticercosis prevalence were origin of the animals (P < 0.001, OR = 7.3) and breeds (P = 0.004, OR = 4.3), and hydatid cysts prevalence was significantly varied with different origins (P = 0.021, OR = 2.8). The viability of C. bovis was higher (28.3%) than that of hydatid cyst (1.7%). Of 79 interviewed respondents, 50.63% had acquired taeniasis and analysis of the risk factors showed association of religions (P = 0.003, OR = 24.4), occupation (P < 0.001, OR = 6.9), educational background (P = 0.035, OR = 2.7) and age (P < 0.001, OR = 3.9) of the respondents with taeniasis prevalence. Furthermore, the inventory of taeniasis drugs dose and treatment cost were estimated to be 335,772 adult doses and 93,310 USD. In conclusion, the findings of the present study imply the zoonotic and socioeconomic importance of the diseases, which need intervention.

Risk factors for foot and mouth disease seroprevalence in indigenous cattle in Southern Ethiopia: the effect of production system.

A serological survey to investigate risk factors for Foot and Mouth Disease (FMD) occurrence was conducted between October 2007 and March 2008 in Southern Ethiopia. Antibodies against non-structural protein of FMD virus (using 3abc ELISA) were measured as indicator of exposure to the virus. The seroprevalence of FMD was 9.5% (95%CI = 7.7 - 11.3, n = 1020) and 48.1% (95% CI = 36.8 - 59.4%, n = 79), respectively at animal and herd levels. Within herd seropositivity was ranged from 6.7 to 46.7% with 18.6% (95%CI = 14.6 - 22.5%) risk of being seropositive for an animal in positive herds. The most important herd level risk factors identified were pastoral system (OR = 16.3, 95% CI = 2.0 -133.7) compared to sedentary, low altitude (OR = 7.5, 95% CI 1.4 -40.7) compared to high altitude, keeping cattle with small ruminants (OR = 5.1, 95% CI 1.0 -25.2) when compared to one species or alone. Seroprevalence was significantly higher (P <0.05) in South Omo than Sidama and Gamo Gofa areas. The odds of seropositivity were 2.8 and 2.3 times higher in the adult (>4 years) and maturing animals (3-4 years) compared to young age category (<3 years). Both multivariable logistic and negative binomial regressions depicted that production system was the major risk factor for FMD seropositivity. Consequently, higher prevalence of FMD in pastoral system where animals are an integral part of life has substantial livelihood and economic implications, which signifies the need for devising control measures.

Survey of tick infestation in small ruminants of Miesso district, West Harergie, Oromia Region, Ethiopia.

A survey was conducted to identify tick species and determine the prevalence of tick infestation in small ruminants of Miesso District, West Harergie Zone. Collection and identification of the ticks were undertaken from November 2007 to April 2008. All visible individual adult ticks were collected from the body of 328 goats and 40 sheep. The prevalence of tick infestation in goats and sheep was found to be 89.9% and 87.5%, respectively. In this study, ten species of ticks which grouped under four genera were identified. The most abundant species found in this study were Boophilus decoloratus (60%), Rhipicephalus pulchellus (25.1%), and Amblyomma gemma (11%). Hyalomma dromedarii was the minor species observed on goats. The difference in the prevalence of tick infestation between sheep and goats was not statistically significant (Chi( 2 ) = 0.22, p = 0.63) but found to be statistically significant between male and females (Chi( 2 ) = 9.8, p = 0.003). Attention should be given to the control and prevention of ticks, since they cause sever damage to the skins of small ruminants and thereby reduce the foreign exchange of the country; they also transmit some diseases which can cause sever loss to the productivity of these animals.

Bovine cysticercosis in cattle slaughtered at Awassa municipal abattoir, Ethiopia: prevalence, cyst viability, distribution and its public health implication.

A cross-sectional study was conducted from October 2005 to April 2006 on bovine cysticercosis in cattle slaughtered at Awassa municipal abattoir with the objective of determining the prevalence of Taenia saginata cysticercosis, cyst viability, distribution and its public health implication. Questionnaire survey involving 120 respondents was also conducted on human taeniasis. A total of 400 carcasses were examined during the study period, of which 105 (26.25%) were infected with T. saginata metacestodes. From a total of 3200 samples inspected, 500 cysticerci were detected in 141 samples, of which 221 (44.2%) were alive. The anatomical distribution of cysticerci were 65 (29.2%) heart, 56 (25.3%) shoulder muscle, 59 (26.7%) masseter, 23 (10.4%) tongue, 12 (5.4%) diaphragm, three (1.4%) liver, two (0.9%) lung and one (0.5%) kidney samples. The prevalence varied significantly between local and crossbred animals (OR = 3.15, P < 0.05), but not varied between sex, age groups and origin of the animals. T. saginata taeniasis was a widespread public health problem in the town with an overall prevalence of 64.2% (77 of 120). The potential risk factors for disease contraction were raw meat consumption, religion and occupational risks. In conclusion, the study revealed high prevalence of T. saginata metacestodes throughout the edible organs together with existence of deep-rooted tradition of raw meat consumption. This may magnify the public health hazards of T. saginata in the study area. As a result, the disease deserves due attention to safeguard the public health and further promote beef industry in the country.

Taeniasis and its socio-economic implication in Awassa town and its surroundings, Southern Ethiopia.

OBJECTIVE: The study was conducted between October 2005 and April 2006 in Awassa town and its surroundings to investigate the status of T. saginata taeniasis, its socio-economic importance and potential risk factors. METHODS: Questionnaire survey was conducted on 120 volunteer individuals selected at random. Additionally, retrospective study on taenicidal drugs inventories at 12 pharmaceutical shops was conducted. RESULTS: T. saginata taeniasis was a wide spread problem in the town and its surroundings with overall prevalence of 64.2% (77 out of 120) and maximum infection frequencies of five times per year. The prevalence of taeniasis was significantly varied between religion, occupation, raw meat consumption and use of spices with raw meat. Accordingly, Christian communities (OR = 5.2, 95% CI = 1.20-22.50), high-risk groups (OR = 4.05, 95% CI = 1.27-12.93), raw meat consumers (OR = 7.9, 95% CI = 2.46-5.66), and spices users (OR = 11.6, 95% CI = 2.85-47.28) had higher likelihood of acquiring taeniasis than Muslim communities, low risk groups, cooked meat eaters and non-spices users, respectively. Conversely, prevalence was not varied between sex, age, marital status and educational backgrounds of the respondents. Respondents' preferences to available taenicidal drugs had strong linear relationship with drug dose sold (R2 = 0.92), and showed that Niclosamide (46.8%) was the drug of first choice while Praziquantel (13%) was the least preferred drug. The taenicidal drug doses and treatment cost from 2002 to 2005 were estimated to be 4,913,346 adult doses and 820,343 USD (7,219,021 Eth. Birr as per rate of 2006). CONCLUSION: Taeniasis was a wide spread problem with higher prevalence and frequent infections in the town and its surroundings. Sociocultural conditions and occupation were the major risk factors for the occurrence of the disease. Hence, owing to its public health and economic importance, taeniasis deserves serious attention in order to safeguard the public health.

Tick infestation of Borana cattle in the Borana Province of Ethiopia.

A study was conducted to identify and determine burdens of ticks infesting Borana cattle in the Borana Province of Ethiopia. Rhipicephalus pulchellus, Rhipicephalus pravus, Rhipicephalus muhsamae, Rhipicephalus praetextatus, Rhipicephalus evertsi evertsi, Amblyomma gemma, Amblyomma variegatum, Amblyomma cohaerens, Amblyomma lepidum, Hyalomma truncatum, Hyalomma marginatum rufipes and Boophilus decoloratus were identified on the cattle. Their burdens ranged from 658-1,554 with a mean of 1,205 ticks per animal. Out of the total 6,025 ticks collected, about 82% were R. pulchellus, followed by R. pravus (about 8%) and A. gemma (5%). The other nine species of ticks accounted for only 5% of the total burdens. It is suggested that a strategic tick control method, aimed mainly at reducing the R. pulchellus burdens, with the objective of allowing a reasonable number of ticks to remain on cattle for the maintenance of endemic stability to tick-borne diseases be instituted.

The use of herbal preparations for tick control in western Ethiopia.

Information on the traditional tick control methods used in Keffa, Illubabor and Wellega Provinces in western Ethiopia was obtained from 86 veterinary clinics and 865 peasant farmers through a questionnaire survey. Latexes of Euphorbia obovalifolia and Ficus brachypoda, juice of crushed leaves of Phytolaca dodecandra and Vernonia amygdalina, fruit juice of Solanum incanum, crushed seeds of Lepidium sativum mixed with fresh cattle faeces, juice of crushed leaves and bark of Calpurnea aurea and commercially available spice of Capsicum spp. mixed with butter, were used by peasant farmers to control ticks. Preliminary in vitro efficacy tests of these plant preparations were performed on engorged female Boophilus decoloratus. Preparations of Capsicum spp., E. obovalifolia, S. incanum and F. brachypoda were found to have 30-100% killing effects. Subsequently, in vivo treatment trials of these preparations were conducted using indigenous Bos indicus cattle naturally infested with ticks. Results indicate that treatments at the rate of once per day for 5 consecutive days with the latexes of E. obovalifolia and F. brachypoda can reduce tick burdens by up to 70% on cattle.

Tick resistance to acaricides in western Ethiopia.

A survey of acaricide resistance was performed on ticks collected from 18 dairy farms and 6 veterinary clinics in western Ethiopia. Boophilus decoloratus collected from most of the dairy farms were resistant to toxaphene, but no resistance was detected in ticks collected from indigenous cattle. There is a need in Ethiopia to replace toxaphene on dairy farms and formulate a rational tick control policy for the whole country.