Molecular cloning, sequencing and recombinant expression of a putative tick protective antigen from three ixodid ticks

The 4D8 gene was recently discovered in Ixodes scapularis and identified as a tick protective antigen. Vaccination using recombinant 4D8 from I. scapularis showed a significant reduction against I. scapularis tick infestation in a sheep model. This protein is expressed in both salivary gland and gut tissues, and is thought to be conserved in ixodid tick species. The objective of this study was to provide evidence of the presence of 4D8 and investigate its sequence homology in three Rhipicephalus tick species from Africa. The gene encoding this tick protective antigen in Rhipicephalus appendiculatus, Rhipicephalus decoloratus and Rhipicephalus microplus ticks was amplified, cloned, sequenced and expressed as a recombinant protein. The amino acid sequences between these three ticks species was found to be conserved with an identity of 96 to 98%. Recombinant 4D8 from the three tick species was expressed as a His-Tag fusion protein in Escherichia coli and the affinity-purified recombinant protein separated by sodium dodecylsulfate polyacrylamide gel electrophoresis (SDS-PAGE), then analyzed in immuno-blot analysis with anti-His-Tag antibody. A unique strong band of the predicted molecular weight of 17 kDa appeared, suggesting presence of a protein corresponding to 4D8. These results confirm the presence of a 4D8 homologue in Rhipicephalus tick stocks from East Africa and further support the hypothesis that it is conserved in different tick species. This conservation among different tick species may suggest that it could potentially be an antigen in subunit vaccines for the control of multiple tick species.Key words: Ticks, vaccine, 4D8, conserved, antigen

Dietary iron status and health of third trimester pregnant women in Kenya: a cross sectional study

Insufficient dietary iron intake is a concern because most foods eaten in developing countries comprise of cereals which are sources of non-heme iron which become bioavailable when eaten with enhancers of absorption. There is an increased demand of iron, particularly in the third trimester, a time an additional supply is required for the growth of foetus and placenta, and to increase the maternal total red cell mass. This study aimed to carry out a comprehensive assessment of the dietary iron intake using 24-hour recall and food frequency questionnaires. Third trimester dietary iron intake survey was a cross-sectional survey of a representative sample of third trimester pregnant women aged 14 – 48 years. Pregnant women were asked to report all foods eaten in the last 24 hours, guided by a glass and asked to state if the quantity of food eaten filled what fraction of the glass. Glass -full of food has a standard mass of 337.5g. The 24 hour recall was repeated for three consecutive days. Quantity of food eaten was compared with the values in Food Consumption Tables to estimate dietary iron intake of each respondent. The bioavailable iron was calculated at 10% of amount of iron in the food for vegetarian populations as recommended in Food Consumption Tables for use in Africa. Mean daily dietary iron intake was 19.62mg (n=109), which was same as the recommended mean of 21mg per day (standard error of the mean 0.6964, Z= -1.02, p<0.05). Furthermore, 15.4% of third trimester pregnant women ate foods with below 19.62mg of iron per. There was no significant correlation between quantity of food eaten and amount of iron in the food (r = 0.0959). The mean daily dietary iron intake is indicative that third trimester pregnant women were mildly iron deficient according to WHO/Office of dietary supplements/Sehmi, and is a suitable index to assess iron status in populations or groups. Key words: dietary iron, iron status, third trimester pregnanc

Bioactivity and toxicity of Bridelia micrantha, Chenopodium ambrosoides and Ocimum americanum plant extracts

Background: Bridelia micrantha, Chenopodium ambrosoides and Ocimum americanum plant species are commonly used in traditional medicine for a number of ailments. The extracts of these plants have been shown to have anti-schistosomal activity suggesting that they could be used for the development of new chemical entities (NCEs) for the treatment of schistosomiasis. However there is limited knowledge on their toxicological profile and their use in traditional medicine may not be a satisfactory safety indication.Methods: In this study the extracts were first screened for bioactivity using brine shrimp lethality test for the determination of LC50 followed by rodent acute toxicity and 28 day subchronic studies.Results: B. micrantha water extract with a LC50 of 77µg/ml was deemed toxic while C. ambrosoides methanol and water extracts were moderately toxic with LC50 of 104.63µg/ml and 696.44µg/ml respectively. O. americanum hexane and water extracts toxicity varied from moderate to slightly toxic with LC50 of 887.59µg/ml and 2254.60µg/ml respectively. C. ambrosoides and O. americanum water extracts which were preferentially selected for subsequent studies were found to have mild to no irritation to rodent eyes and skin. Moreover, the aminotransferases AST and ALT which were used to detect liver injury suggested negligible effect.Conclusions: This therefore confirms that C. ambrosoides and O. americanum water extracts are safe for clinical use with O. americanum water extract having a slight edge

Progress in achieving household food security in climate-smart villages in the Albertine Rift, western Uganda

Hoima is located in western Uganda east of Lake Albert, on a landscape that is generally undulating with relatively flat low lying area alternating with broad hills. The area has a population density of 160 persons per square kilometer, with 22% of the people living below the poverty line. The area faces land degradation and declining soil fertility. The key food crops are cassava, beans, sweet potatoes, and maize. Chicken, pigs, cows and goats are important for food and income generation. Most households get their food supplies from their own farms throughout the year. The worst months for food supplies, when more than 20% of households get their food mainly from off-farm sources and 40% of the households have food deficits are March and April, which also mark the beginning of the rains after several months of dry season. About 31% of households are food secure all year long. Another 35% suffer food deficits for 1-2 months per year. 16% of these households struggle to get enough to feed their families for 3-4 months, 9% for 5-6 months, and 10% for more than six months per year

Unique Mitochondrial Single Nucleotide Polymorphisms Demonstrate Resolution Potential to Discriminate Theileria parva Vaccine and Buffalo-Derived Strains

Distinct pathogenic and epidemiological features underlie different Theileria parva strains resulting in different clinical manifestations of East Coast Fever and Corridor Disease in susceptible cattle. Unclear delineation of these strains limits the control of these diseases in endemic areas. Hence, an accurate characterization of strains can improve the treatment and prevention approaches as well as investigate their origin. Here, we describe a set of single nucleotide polymorphisms (SNPs) based on 13 near-complete mitogenomes of T. parva strains originating from East and Southern Africa, including the live vaccine stock strains. We identified 11 SNPs that are non-preferentially distributed within the coding and non-coding regions, all of which are synonymous except for two within the cytochrome b gene of buffalo-derived strains. Our analysis ascertains haplotype-specific mutations that segregate the different vaccine and the buffalo-derived strains except T. parva-Muguga and Serengeti-transformed strains suggesting a shared lineage between the latter two vaccine strains. Phylogenetic analyses including the mitogenomes of other Theileria species: T. annulata, T. taurotragi, and T. lestoquardi, with the latter two sequenced in this study for the first time, were congruent with nuclear-encoded genes. Importantly, we describe seven T. parva haplotypes characterized by synonymous SNPs and parsimony-informative characters with the other three transforming species mitogenomes. We anticipate that tracking T. parva mitochondrial haplotypes from this study will provide insight into the parasite’s epidemiological dynamics and underpin current control efforts

Evidence for conservation in antigen gene sequences combined with extensive polymorphism at VNTR loci

Theileria parva is a tick‐transmitted apicomplexan protozoan parasite that infects lymphocytes of cattle and African Cape buffalo (Syncerus caffer), causing a frequently fatal disease of cattle in eastern, central and southern Africa. A live vaccination procedure, known as infection and treatment method (ITM), the most frequently used version of which comprises the Muguga, Serengeti‐transformed and Kiambu 5 stocks of T. parva, delivered as a trivalent cocktail, is generally effective. However, it does not always induce 100% protection against heterologous parasite challenge. Knowledge of the genetic diversity of T. parva in target cattle populations is therefore important prior to extensive vaccine deployment. This study investigated the extent of genetic diversity within T. parva field isolates derived from Ankole (Bos taurus) cattle in south‐western Uganda using 14 variable number tandem repeat (VNTR) satellite loci and the sequences of two antigen‐encoding genes that are targets of CD8+T‐cell responses induced by ITM, designated Tp1 and Tp2. The findings revealed a T. parva prevalence of 51% confirming endemicity of the parasite in south‐western Uganda. Cattle‐derived T. parva VNTR genotypes revealed a high degree of polymorphism. However, all of the T. parva Tp1 and Tp2 alleles identified in this study have been reported previously, indicating that they are widespread geographically in East Africa and highly conserved

In Vitro Gas, Methane, and Carbon Dioxide Productions of High Fibrous Diet Incubated With Fecal Inocula From Horses in Response to the Supplementation With Different Live Yeast Additives

Yeast supplementation of horse diets can influence nutrient digestibility and microbiota dynamics in the horse hindgut. In some in vitro [6] and in vivo [4] studies, yeast addition to the diets improved digestion of low-quality forages. It has been shown that yeast supplementation can alter the microbial environment by increasing the total number of hindgut microorganisms [7]. As a result, feed digestion in the hindgut can be enhanced, especially that of the fiber fraction, most likely due to increased numbers of cellulolytic bacteria in the hindgut [8]. In contrast, other studies have reported no effect of yeast addition to equine diets on nutrient digestibility in vitro [7] or in vivo [9].In a randomized block design experiment, the effect of fecal inocula from horses supplemented with live yeast (Saccharomyces cerevisiae) in diets containing 50% oat straw on in vitro total gas (gas production [GP]), methane (CH4), and carbon dioxide (CO2) productions as indicators of hindgut activity was assessed. Three commercial products of S. cerevisiae were tested (1) Biocell F53 (YST53), (2) Procreatin 7 (YST07), and (3) Biosaf SC47 (YST047). For the incubations, each product was added at 0 (control without yeast addition), 2, or 4 mg/g dry matter (DM). Fecal inocula for incubations with each treatment was obtained from Quarter Horse mares fed the same yeast additives for 15 days, resulting in four different fecal inocula (FI53, FI07, FI47, and FI00). The fecal content mixed with the culture media were used to inoculate three identical runs of incubation in bottles containing 1-g DM of substrate (a mixture of concentrate and oat straw [1:1 DM]). The GP, CH4, and CO2 productions were measured at 2, 4, 6, 8, 10, 12, 24, and 48 hours postincubation. Addition of additives YST53 and YST07 at 2 mg/g DM resulted in higher asymptotic GP (linear effect, P ¼.021) and GP during the first 12 hours of incubation (linear effect, P <.05) compared with control without yeast addition, with the highest value being for the dose 2 mg/g DM with the fecal inoculum FI53. The additive YST47 at all doses with fecal inoculum FI47 had lower GP (linear effect, P < .05) at different incubation hours compared with control. The additive YST53 increased GP, CH4, and fermentation kinetics at the dose 2 mg/g DM with decreasing CH4 production by 78% at 4 mg/g DM at 24 hours of incubation. Addition of YST53 at 2 and 4 mg/g DM with fecal inoculum FI53 enhanced fermentation kinetics (P < .05) compared with control and other additives at different doses. It can be concluded that the yeast additive Biocell F53 was the most effective at doses of 2 and 4 mg/g DM compared with other Saccharomyces strains to attain a more favorable hindgut fermentation to digest fibrous roughages by horses

Limited diversity in the CD8+ antigen-coding loci in Theileria parva parasites from cattle from southern and eastern Africa

Theileria parva infections in cattle causes huge economic losses in the affected African countries, directly impacting the livelihood of the poor small-holder farmers. The current immunization protocol using live sporozoites in eastern Africa, is among the control measures designed to limit T. parva infections in cattle. However, the ability of the immune protection induced by this immunization to protect against field parasites has been compromised by the diversity of the parasite involving the schizont antigen genes. Previous studies have reported on the antigenic diversity of T. parva parasites from southern and eastern Africa, however, similar reports on T. parva parasites particularly from cattle from southern Africa remains scanty, due to the self-limiting nature of Corridor disease. Thus, we evaluated the diversity of CD8+ T-cell regions of ten schizont antigen genes in T. parva parasites associated with Corridor disease and East Coast fever (ECF) from southern and eastern Africa respectively. Regions of schizont antigen (TpAg) genes containing the CD8+ T-cell epitopes (CTL determinants) were amplified from genomic DNA extracted from blood of T. parva positive samples, cloned and sequenced. The results revealed limited diversity between the two parasite groups from cattle from southern and eastern Africa, defying the widely accepted notion that antigen-encoding loci in cattle-derived parasites are conserved, while in buffalo-derived parasites, they are extensively variable. This suggests that only a sub-population of parasites is successfully transmitted from buffalo to cattle, resulting in the limited antigenic diversity in Corridor disease parasites. Tp4, Tp5, Tp7 and Tp8 showed limited to absence of diversity in both parasite groups, suggesting the need to further investigate their immunogenic properties for consideration as candidates for a subunit vaccine. Distinct and common variants of Tp2 were detected among the ECF parasites from eastern Africa indicating evidence of parasite mixing following immunization. This study provides additional information on the comparative diversity of TpAg genes in buffalo- and cattle-derived T. parva parasites from cattle from southern and eastern Africa.The National Research Foundation, South Africa; University of Pretoria, South Africa; and the National Research Fund, Kenya.https://www.elsevier.com/locate/vetparhj2022Veterinary Tropical Disease