Options for selecting dairy cattle for milk coagulation ability

Lypsylehmien maidon juoksettumiskyvyn jalostuskeinot Väitöskirjassa tutkittiin lypsylehmien maidon juustonvalmistuslaadun parantamista jalostusvalinnan avulla. Tutkimusaihe on tärkeä, sillä yhä suurempi osa maidosta käytetään juustonvalmistukseen. Tutkimuksen kohteena oli maidon juoksettumiskyky, sillä se on yksi keskeisistä juustomäärään vaikuttavista tekijöistä. Maidon juoksettumiskyky vaihteli huomattavasti lehmien, sonnien, karjojen, rotujen ja lypsykauden vaiheiden välillä. Vaikka tankkimaidon juoksettumiskyvyssä olikin suuria eroja karjoittain, karja selitti vain pienen osan juoksettumiskyvyn kokonaisvaihtelusta. Todennäköisesti perinnölliset erot lehmien välillä selittävät suurimman osan karjojen tankkimaitojen juoksettumiskyvyssä havaituista eroista. Hyvä hoito ja ruokinta vähensivät kuitenkin jossain määrin huonosti juoksettuvien tankkimaitojen osuutta karjoissa. Holstein-friisiläiset lehmät olivat juoksettumiskyvyltään ayrshire-rotuisia lehmiä parempia. Huono juoksettuminen ja juoksettumattomuus oli vain vähäinen ongelma holstein-friisiläisillä (10 %), kun taas kolmannes ayrshire-lehmistä tuotti huonosti juoksettuvaa tai juoksettumatonta maitoa. Maitoa sanotaan huonosti juoksettuvaksi silloin, kun juustomassa ei ole riittävän kiinteää leikattavaksi puolen tunnin kuluttua juoksetteen lisäyksestä. Juoksettumattomaksi määriteltävä maito ei saostu lainkaan puolen tunnin aikana ja on siksi erittäin huonoa raaka-ainetta juustomeijereille. Noin 40 % lehmien välisistä eroista maidon juoksettumiskyvyssä selittyi perinnöllisillä tekijöillä. Juoksettumiskykyä voikin sanoa hyvin periytyväksi ominaisuudeksi. Kolme mittauskertaa lehmää kohti riittää varsin hyvin lehmän maidon keskimääräisen juoksettumiskyvyn arvioimiseen. Tällä hetkellä juoksettumiskyvyn suoran jalostamisen ongelmana on kuitenkin automatisoidun, laajamittaiseen käyttöön soveltuvan mittalaitteen puute. Tämän takia väitöskirjassa tutkittiin mahdollisuuksia jalostaa maidon juoksettumiskykyä epäsuorasti, jonkin toisen ominaisuuden kautta. Tällaisen ominaisuuden pitää olla kyllin voimakkaasti perinnöllisesti kytkeytynyt juoksettumiskykyyn, jotta jalostus olisi mahdollista sen avulla. Tutkittavat ominaisuudet olivat sonnien kokonaisjalostusarvossa jo mukana olevat maitotuotos ja utareterveyteen liittyvät ominaisuudet sekä kokonaisjalostusarvoon kuulumattomat maidon valkuais- ja kaseiinipitoisuus sekä maidon pH. Väitöskirjassa tutkittiin myös mahdollisuuksia ns. merkkiavusteiseen valintaan tutkimalla maidon juoksettumattomuuden perinnöllisyyttä ja kartoittamalla siihen liittyvät kromosomialueet. Tutkimuksen tulosten perusteella lehmien utareterveyden jalostaminen parantaa jonkin verran myös maidon juoksettumiskykyä sekä vähentää juoksettumattomuutta ayrshire-rotuisilla lehmillä. Lehmien maitotuotos ja maidon juoksettumiskyky sekä juoksettumattomuus ovat sen sijaan perinnöllisesti toisistaan riippumattomia ominaisuuksia. Myöskin maidon valkuais- ja kaseiinipitoisuuden perinnöllinen yhteys juoksettumiskykyyn oli likimain nolla. Maidon pH:n ja juoksettumiskyvyn välillä oli melko voimakas perinnöllinen yhteys, joten maidon pH:n jalostaminen parantaisi myös maidon juoksettumiskykyä. Todennäköisesti sen jalostaminen ei kuitenkaan vähentäisi juoksettumatonta maitoa tuottavien lehmien määrää. Koska maidon juoksettumattomuus on niin yleinen ongelma suomalaisilla ayrshire-lehmillä, väitöksessä selvitettiin tarkemmin ilmiön taustoja. Kaikissa kolmessa tutkimusaineistoissa noin 10 % ayrshire-lehmistä tuotti juoksettumatonta maitoa. Kahden vuoden kuukausittaisen seurannan aikana osa lehmistä tuotti juoksettumatonta maitoa lähes joka mittauskerralla. Maidon juoksettumattomuus oli yhteydessä lypsykauden vaiheeseen, mutta mikään ympäristötekijöistä ei pystynyt täysin selittämään sitä. Sen sijaan viitteet sen periytyvyydestä vahvistuivat tutkimusten edetessä. Lopuksi tutkimusryhmä onnistui kartoittamaan juoksettumattomuutta aiheuttavat kromosomialueet kromosomeihin 2 ja 18, lähelle DNA-merkkejä BMS1126 ja BMS1355. Tulosten perusteella maidon juoksettumattomuus ei ole yhteydessä maidon juoksettumistapahtumassa keskeisiin kaseiinigeeneihin. Sen sijaan on mahdollista, että juoksettumattomuusongelman aiheuttavat kaseiinigeenien syntetisoinnin jälkeisessä muokkauksessa tapahtuvat virheet. Asia vaatii kuitenkin perusteellista tutkimista. Väitöksen tulosten perusteella maidon juoksettumattomuusgeeniä kantavien eläinten karsiminen jalostuseläinten joukosta olisi tehokkain tapa jalostaa maidon juoksettumiskykyä suomalaisessa lypsykarjapopulaatiossa.

Quantitative trait loci detection and benefits from marker-assisted selection in dairy cattle

Knowing the chromosomal areas or actual genes affecting the traits under selection would add more information to be used in the selection decisions which would potentially lead to higher genetic response. The first objective of this study was to map quantitative trait loci (QTL) affecting economically important traits in the Finnish Ayrshire population. The second objective was to investigate the effects of using QTL information in marker-assisted selection (MAS) on the genetic response and the linkage disequilibrium between the different parts of the genome. Whole genome scans were carried out on a grand-daughter design with 12 half-sib families and a total of 493 sons. Twelve different traits were studied: milk yield, protein yield, protein content, fat yield, fat content, somatic cell score (SCS), mastitis treatments, other veterinary treatments, days open, fertility treatments, non-return rate, and calf mortality. The average spacing of the typed markers was 20 cM with 2 to 14 markers per chromosome. Associations between markers and traits were analyzed with multiple marker regression. Significance was determined by permutation and genome-wise P-values obtained by Bonferroni correction. The benefits from MAS were investigated by simulation: a conventional progeny testing scheme was compared to a scheme where QTL information was used within families to select among full-sibs in the male path. Two QTL on different chromosomes were modelled. The effects of different starting frequencies of the favourable alleles and different size of the QTL effects were evaluated. A large number of QTL, 48 in total, were detected at 5% or higher chromosome-wise significance. QTL for milk production were found on 8 chromosomes, for SCS on 6, for mastitis treatments on 1, for other veterinary treatments on 5, for days open on 7, for fertility treatments on 7, for calf mortality on 6, and for non-return rate on 2 chromosomes. In the simulation study the total genetic response was faster with MAS than with conventional selection and the advantage of MAS persisted over the studied generations. The rate of response and the difference between the selection schemes reflected clearly the changes in allele frequencies of the favourable QTL. The disequilibrium between the polygenes and QTL was always negative and it was larger with larger QTL size. The disequilibrium between the two QTL was larger with QTL of large effect and it was somewhat larger with MAS for scenarios with starting frequencies below 0.5 for QTL of moderate size and below 0.3 for large QTL. In conclusion, several QTL affecting economically important traits of dairy cattle were detected. Further studies are needed to verify these QTL, check their presence in the present breeding population, look for pleiotropy and fine map the most interesting QTL regions. The results of the simulation studies show that using MAS together with embryo transfer to pre-select young bulls within families is a useful approach to increase the genetic merit of the AI-bulls compared to conventional selection.

Ammonia emissions from pig and cattle slurry in the field and utilization of slurry nitrogen in crop production

Volatilization of ammonia (NH3) from animal manure is a major pathway for nitrogen (N) losses that cause eutrophication, acidification, and other environmental hazards. In this study, the effect of alternative techniques of manure treatment (aeration, separation, addition of peat) and application (broadcast spreading, band spreading, injection, incorporation by harrowing) on ammonia emissions in the field and on nitrogen uptake by ley or cereals was studied. The effect of a mixture of slurry and peat on soil properties was also investigated. The aim of this study was to find ways to improve the utilization of manure nitrogen and reduce its release to the environment. Injection into the soil or incorporation by harrowing clearly reduced ammonia volatilization from slurry more than did the surface application onto a smaller area by band spreading or reduction of the dry matter of slurry by aeration or separation. Surface application showed low ammonia volatilization, when pig slurry was applied to tilled bare clay soil or to spring wheat stands in early growth stages. Apparently, the properties of both slurry and soil enabled the rapid infiltration and absorption of slurry and its ammoniacal nitrogen by the soil. On ley, however, surface-applied cattle slurry lost about half of its ammoniacal nitrogen. The volatilization of ammonia from surface-applied peat manure was slow, but proceeded over a long period of time. After rain or irrigation, the peat manure layer on the soil surface retarded evaporation. Incorporation was less important for the fertilizer effect of peat manure than for pig slurry, but both manures were more effective when incorporated. Peat manure applications increase soil organic matter content and aggregate stability. Stubble mulch tillage hastens the effect in surface soil compared with ploughing. The apparent recovery of ammoniacal manure nitrogen in crop yield was higher with injection and incorporation than with surface applications. This was the case for leys as well as for spring cereals, even though ammonia losses from manures applied to cereals were relatively low with surface applications as well. The ammoniacal nitrogen of surface-applied slurry was obviously adsorbed by the very surface soil and remained mostly unavailable to plant roots in the dry soil. Supplementing manures with inorganic fertilizer nitrogen, which adds plant-available nitrogen to the soil at the start of growth, increased the overall recovery of applied nitrogen in crop yields.

Maintaining plant species richness by cattle grazing - mesic semi-natural grasslands as focal habitats

Semi-natural grasslands are the most important agricultural areas for biodiversity. The present study investigates the effects of traditional livestock grazing and mowing on plant species richness, the main emphasis being on cattle grazing in mesic semi-natural grasslands. The two reviews provide a thorough assessment of the multifaceted impacts and importance of grazing and mowing management to plant species richness. It is emphasized that livestock grazing and mowing have partially compensated the suppression of major natural disturbances by humans and mitigated the negative effects of eutrophication. This hypothesis has important consequences for nature conservation: A large proportion of European species originally adapted to natural disturbances may be at present dependent on livestock grazing and / or mowing. Furthermore, grazing and mowing are key management methods to mitigate effects of nutrient-enrichment. The species composition and richness in old (continuously grazed), new (grazing restarting 3-8 years ago) and abandoned (over 10 years) pastures differed consistently across a range of spatial scales, and was intermediate in new pastures compared to old and abandoned pastures. In mesic grasslands most plant species were shown to benefit from cattle grazing. Indicator species of biologically valuable grasslands and rare species were more abundant in grazed than in abandoned grasslands. Steep S-SW-facing slopes are the most suitable sites for many grassland plants and should be prioritized in grassland restoration. The proportion of species trait groups benefiting from grazing was higher in mesic semi-natural grasslands than in dry and wet grasslands. Consequently, species trait responses to grazing and the effectiveness of the natural factors limiting plant growth may be intimately linked High plant species richness of traditionally mowed and grazed areas is explained by numerous factors which operate on different spatial scales. Particularly important for maintaining large scale plant species richness are evolutionary and mitigation factors. Grazing and mowing cause a shift towards the conditions that have occurred during the evolutionary history of European plant species by modifying key ecological factors (nutrients, pH and light). The results of this Dissertation suggest that restoration of semi-natural grasslands by private farmers is potentially a useful method to manage biodiversity in the agricultural landscape. However, the quality of management is commonly improper, particularly due to financial constraints. For enhanced success of restoration, management regulations in the agri-environment scheme need to be defined more explicitly and the scheme should be revised to encourage management of biodiversity.

Habitat use and population dynamics of brown bears (Ursus arctos) in Scandinavia

Large carnivore populations are currently recovering from past extirpation efforts and expanding back into their original habitats. At the same time human activities have resulted in very few wilderness areas left with suitable habitats and size large enough to maintain populations of large carnivores without human contact. Consequently the long-term future of large carnivores depends on their successful integration into landscapes where humans live. Thus, understanding their behaviour and interaction with surrounding habitats is of utmost importance in the development of management strategies for large carnivores. This applies also to brown bears (Ursus arctos) that were almost exterminated from Scandinavia and Finland at the turn of the century, but are now expanding their range with the current population estimates being approximately 2600 bears in Scandinavia and 840 in Finland. This thesis focuses on the large-scale habitat use and population dynamics of brown bears in Scandinavia with the objective to develop modelling approaches that support the management of bear populations. Habitat analysis shows that bear home ranges occur mainly in forested areas with a low level of human influence relative to surrounding areas. Habitat modelling based on these findings allows identification and quantification of the potentially suitable areas for bears in Scandinavia. Additionally, this thesis presents novel improvements to home range estimation that enable realistic estimates of the effective area required for the bears to establish a home range. This is achieved through fitting to the radio-tracking data to establish the amount of temporal autocorrelation and the proportion of time spent in different habitat types. Together these form a basis for the landscape-level management of the expanding population. Successful management of bears requires also assessment of the consequences of harvest on the population viability. An individual-based simulation model, accounting for the sexually selected infanticide, was used to investigate the possibility of increasing the harvest using different hunting strategies, such as trophy harvest of males. The results indicated that the population can sustain twice the current harvest rate. However, harvest should be changed gradually while carefully monitoring the population growth as some effects of increased harvest may manifest themselves only after a time-delay. The results and methodological improvements in this thesis can be applied to the Finnish bear population and to other large carnivores. They provide grounds for the further development of spatially-realistic management-oriented models of brow bear dynamics that can make projections of the future distribution of bears while accounting for the development of human activities.

Species-specific and Indication-based Use of Antimicrobials in Dogs, Cats, Cattle and Horses in Finland : Data collected using three different methods

Increasing antimicrobial resistance in bacteria has led to the need for better understanding of antimicrobial usage patterns. In 1999, the World Organisation for Animal Health (OIE) recommended that an international ad hoc group should be established to address human and animal health risks related to antimicrobial resistance and the contribution of antimicrobial usage in veterinary medicine. In European countries the need for continuous recording of the usage of veterinary antimicrobials as well as for animal species-specific and indication-based data on usage has been acknowledged. Finland has been among the first countries to develop prudent use guidelines in veterinary medicine, as the Ministry of Agriculture and Forestry issued the first animal species-specific indication-based recommendations for antimicrobial use in animals in 1996. These guidelines have been revised in 2003 and 2009. However, surveillance on the species-specific use of antimicrobials in animals has not been performed in Finland. This thesis provides animal species-specific information on indication-based antimicrobial usage. Different methods for data collection have been utilized. Information on antimicrobial usage in animals has been gathered in four studies (studies A-D). Material from studies A, B and C have been used in an overlapping manner in the original publications I-IV. Study A (original publications I & IV) presents a retrospective cross-sectional survey on prescriptions for small animals at the Veterinary Teaching Hospital of the University of Helsinki. Prescriptions for antimicrobial agents (n = 2281) were collected and usage patterns, such as the indication and length of treatment, were reviewed. Most of the prescriptions were for dogs (78%), and primarily for the treatment of skin and ear infections most of which were treated with cephalexin for a median period of 14 days. Prescriptions for cats (18%) were most often for the treatment of urinary tract infections with amoxicillin for a median length of 10 days. Study B (original publication II) was a retrospective cross-sectional survey where prescriptions for animals were collected from 17 University Pharmacies nationwide. Antimicrobial prescriptions (n = 1038) for mainly dogs (65%) and cats (19%) were investigated. In this study, cephalexin and amoxicillin were also the most frequently used drugs for dogs and cats, respectively. In study C (original publications III & IV), the indication-based usage of antimicrobials of practicing veterinarians was analyzed by using a prospective questionnaire. Randomly selected practicing veterinarians in Finland (n = 262) recorded all their antimicrobial usage during a 7-day study period. Cattle (46%) with mastitis were the most common patients receiving antimicrobial treatment, generally intramuscular penicillin G or intramammary treatment with ampicillin and cloxacillin. The median length of treatment was four days, regardless of the route of administration. Antimicrobial use in horses was evaluated in study D, the results of which are previously unpublished. Firstly, data collected with the prospective questionnaire from the practicing veterinarians showed that horses (n = 89) were frequently treated for skin or wound infections by using penicillin G or trimethoprim-sulfadiazine. The mean duration of treatment was five to seven days. Secondly, according to retrospective data collected from patient records, horses (n = 74) that underwent colic surgery at the Veterinary Teaching Hospital of the University of Helsinki were generally treated according to national and hospital recommendations; penicillin G and gentamicin was administered preoperatively and treatment was continued for a median of three days postoperatively. In conclusion, Finnish veterinarians followed well the national prudent use guidelines. Narrow-spectrum antimicrobials were preferred and, for instance, fluoroquinolones were used sparingly. Prescription studies seemed to give good information on antimicrobials usage, especially when combined with complementary information from patient records. A prospective questionnaire study provided a fair amount of valuable data on several animal species. Electronic surveys are worthwhile exploiting in the future.

Finnish cattle as reservoir of Campylobacter spp.

The reported incidence of human campylobacteriosis in Finland is higher than in most other European countries. A high annual percentage of sporadic infections is of foreign origin, although a notable proportion of summer infections is domestically acquired. While chickens appear to be a major source of campylobacters for humans in most countries, the prevalence of campylobacters is very low in chicken slaughter batches in Finland. Data on other potential animal reservoirs of human pathogenic campylobacters in Finland are scarce. Consequently, this study aimed to investigate the status of Finnish cattle as a potential source of thermophilic Campylobacter spp. and antibiotic-resistant Campylobacter jejuni for human sporadic campylobacter infections of domestic origin. A survey of the prevalence of thermophilic Campylobacter spp. in Finnish cattle studied bovine rectal faecal samples (n=952) and carcass surface samples (n=948) from twelve Finnish slaughterhouses from January to December 2003. The total prevalence of Campylobacter spp. in faecal samples was 31.1%, and in carcass samples 3.5%. Campylobacter jejuni, the most common species, was present in 19.5% of faecal samples and in 3.1% of carcasses. In addition to thermophilic Campylobacter spp., C. hyointestinalis ssp. hyointestinalis was present in bovine samples. The prevalence of campylobacters was higher among beef cattle than among dairy cattle. Using the enrichment method, the number of positive faecal samples was 7.5 times higher than that obtained by direct plating. The predominant serotypes of faecal C. jejuni, determined by serotyping with a set of 25 commercial antisera for heat-stable antigens (Penner), were Pen2 and Pen4-complex, which covered 52% of the samples. Genotyping with pulsed-field gel electrophoresis (PFGE) using SmaI restriction yielded a high diversity of C. jejuni subtypes in cattle. Determining the minimum inhibitory concentrations of ampicillin, enrofloxacin, erythromycin, gentamicin, nalidixic acid, and oxytetracycline among bovine C. jejuni isolates using a commercial broth microdilution method yielded 9% of isolates resistant to at least one of the antimicrobials examined. No multiresistant isolates were found among the bovine C. jejuni strains. The study of the shedding patterns of Campylobacter spp. among three Finnish dairy cattle herds included the examination of fresh faecal samples and tank milk samples taken five times, as well as samples from drinking troughs taken once during the one-year study. The semiquantitative enrichment method detected C. jejuni in 169 of the 340 faecal samples, mostly at low levels. In addition, C. jejuni was present in one drinking trough sample. The prevalence between herds and sampling occasions varied widely. PFGE, using SmaI as restriction enzyme, identified only a few subtypes in each herd. In two 2 of the herds, two subtypes persisted throughout the sampling. Individual animals presented various shedding patterns during the study. Comparison of C. jejuni isolates from humans, chickens and cattle included the design of primers for four new genetic markers selected from completely sequenced C. jejuni genomes 81-176, RM1221 and NCTC 11168, and the PCR examination of domestic human isolates from southern Finland in 1996, 2002 and 2003 (n=309), chicken isolates from 2003, 2006 and 2007 (n=205), and bovine isolates from 2003 (n=131). The results revealed that bovine isolates differed significantly from human and chicken isolates. In particular, the - glutamyl transpeptidase gene was uncommon among bovine isolates. The PFGE genotyping of C. jejuni isolates, using SmaI and KpnI restriction enzymes, included a geographically representative collection of isolates from domestic sporadic human infections, chicken slaughter batches, and cattle faeces and carcasses during the seasonal peak of campylobacteriosis in the summer of 2003. The study determined that 55.4% of human isolates were indistinguishable from those of chickens and cattle. Temporal association between isolates from humans and chickens was possible in 31.4% of human infections. Approximately 19% of the human infections may have been associated with cattle. However, isolates from bovine carcasses and human cases represented different PFGE subtypes. In conclusion, this study suggests that Finnish cattle is a notable reservoir of C. jejuni, the most important Campylobacter sp. in human enteric infections. Although the concentration of these organisms in bovine faeces appeared to be low, excretion can be persistent. The genetic diversity and presence or absence of marker genes support previous suggestions of host-adapted C. jejuni strains, and may indicate variations in virulence between strains from different hosts. In addition to chickens, Finnish cattle appeared to be an important reservoir and possible source of C. jejuni in domestic sporadic human infections. However, sources of campylobacters may differ between rural and urban areas in Finland, and in general, the transmission of C. jejuni of bovine origin probably occurs via other routes than food.

Cattle Teeth in Graves : Interpretations of animal bones found in Finnish inhumation graves (ca AD 550-1700)

The nature of a burial is always ritualistic. This is often forgotten when dealing with Finnish inhumation burials containing animal bones. Only the animal bones found close to the deceased have traditionally been thought to have a ritualistic purpose. The animal bones found in the filling of the grave, which is still part of the burial, has on the other hand, often been neglected in the previous research. In this Master s thesis I will discuss the function and interpretation of animal bones in graves. The base of this study is six sites, all of different nature, from Finland. Luistari in Eura is from the western coast and is dated to Late Iron Age (and possibly Medieval period), the Medieval hamlet of Finno is situated in Espoo which is situated on the southern coast. Two town burials, Turku and Porvoo, are also included in this study. The graves from Turku are dated to Late Medieval period and Early Renaissance, whereas the cemetery in Porvoo is from the 18th century. Visulahti in Mikkeli is from the Late Iron Age and represents Eastern Finnish burial tradition, the same as Suotniemi from Käkisalmi parish, which is nowadays part of Russia. While parts of the animal bones had already been analysed before, the author also analysed animal bones for the purpose of the present Master´s thesis. The bones were compared to the burial contexts, when possible. Based on the comparisons I have made interpretations which might explain the existence of animal bones in the graves. The interpretations are among others sacrifice, commemoration meals and animal burials. The site could also have been a settlement site prior to the graves, thus the bones in the graves would belong to the settlement phase. When comparing the date of the studied sites, the town burials are later and the animal bones are probably related to previous or contemporary use of the sites as graveyards. On top of this there does not seem to be much difference in burial tradition between Eastern and Western Finland, although at least from the hamlet burials of Finno there are aspects that could be linked to Eastern burials. In making the interpretations I have taken into consideration the aspects of belief during different time periods when they could be accounted as relevant. Also the problems with bone preservation were relevant and challenging for the study. Often only the hardest substance of the skeleton, namely teeth, has been preserved. For this reason the quality of the archaeological documentation was a key issue in this study. In producing quality interpretations of the animal bones in graves, the bones, contexts and their relationship to the surrounding site should be documented with care.

Measurement of acceleration while walking as an automated method for gait assessment in dairy cattle

The aims were to determine whether measures of acceleration of the legs and back of dairy cows while they walk could help detect changes in gait or locomotion associated with lameness and differences in the walking surface. In 2 experiments, 12 or 24 multiparous dairy cows were fitted with five 3-dimensional accelerometers, 1 attached to each leg and 1 to the back, and acceleration data were collected while cows walked in a straight line on concrete (experiment 1) or on both concrete and rubber (experiment 2). Cows were video-recorded while walking to assess overall gait, asymmetry of the steps, and walking speed. In experiment 1, cows were selected to maximize the range of gait scores, whereas no clinically lame cows were enrolled in experiment 2. For each accelerometer location, overall acceleration was calculated as the magnitude of the 3-dimensional acceleration vector and the variance of overall acceleration, as well as the asymmetry of variance of acceleration within the front and rear pair of legs. In experiment 1, the asymmetry of variance of acceleration in the front and rear legs was positively correlated with overall gait and the visually assessed asymmetry of the steps (r ≥0.6). Walking speed was negatively correlated with the asymmetry of variance of the rear legs (r=−0.8) and positively correlated with the acceleration and the variance of acceleration of each leg and back (r ≥0.7). In experiment 2, cows had lower gait scores [2.3 vs. 2.6; standard error of the difference (SED)=0.1, measured on a 5-point scale] and lower scores for asymmetry of the steps (18.0 vs. 23.1; SED=2.2, measured on a continuous 100-unit scale) when they walked on rubber compared with concrete, and their walking speed increased (1.28 vs. 1.22m/s; SED=0.02). The acceleration of the front (1.67 vs. 1.72g; SED=0.02) and rear (1.62 vs. 1.67g; SED=0.02) legs and the variance of acceleration of the rear legs (0.88 vs. 0.94g; SED=0.03) were lower when cows walked on rubber compared with concrete. Despite the improvements in gait score that occurred when cows walked on rubber, the asymmetry of variance of acceleration of the front leg was higher (15.2 vs. 10.4%; SED=2.0). The difference in walking speed between concrete and rubber correlated with the difference in the mean acceleration and the difference in the variance of acceleration of the legs and back (r ≥0.6). Three-dimensional accelerometers seem to be a promising tool for lameness detection on farm and to study walking surfaces, especially when attached to a leg.

Molecular Characterization of Viruses Causing the Cassava Brown Streak Disease Epidemic in Eastern Africa

Cassava brown streak disease (CBSD) was described for the first time in Tanganyika (now Tanzania) about seven decades ago. Tanganyika (now Tanzania) about seven decades ago. It was endemic in the lowland areas of East Africa and inland parts of Malawi and caused by Cassava brown streak virus (CBSV; genus Ipomovirus; Potyviridae). However, in 1990s CBSD was observed at high altitude areas in Uganda. The causes for spread to new locations were not known.The present work was thus initiated to generate information on genetic variability, clarify the taxonomy of the virus or viruses associated with CBSD in Eastern Africa as well as to understand the evolutionary forces acting on their genes. It also sought to develop a molecular based diagnostic tool for detection of CBSD-associated virus isolates. Comparison of the CP-encoding sequences of CBSD-associated virus isolates collected from Uganda and north-western Tanzania in 2007 and the partial sequences available in Genbank revealed occurrence of two genetically distinct groups of isolates. Two isolates were selected to represent the two groups. The complete genomes of isolates MLB3 (TZ:Mlb3:07) and Kor6 (TZ:Kor6:08) obtained from North-Western (Kagera) and North-Eastern (Tanga) Tanzania, respectively, were sequenced. The genomes were 9069 and 8995 nucleotides (nt), respectively. They translated into polyproteins that were predicted to yield ten mature proteins after cleavage. Nine proteins were typical in the family Potyviridae, namely P1, P3, 6K1, CI, 6K2, VPg, NIa-Pro, NIb and CP, but the viruses did not contain HC-Pro. Interestingly, genomes of both isolates contained a Maf/HAM1-like sequence (HAM1h; 678 nucleotides, 25 kDa) recombined between the NIb and CP domains in the 3’-proximal part of the genomes. HAM1h was also identified in Euphorbia ringspot virus (EuRSV) whose sequence was in GenBank. The HAM1 gene is widely spread in both prokaryotes and eukaryotes. In yeast (Saccharomyces cerevisiae) it is known to be a nucleoside triphosphate (NTP) pyrophosphatase. Novel information was obtained on the structural variation at the N-termini of polyproteins of viruses in the genus Ipomovirus. Cucumber vein yellowing virus (CVYV) and Squash vein yellowing virus (SqVYV) contain a duplicated P1 (P1a and P1b) but lack the HC-Pro. On the other hand, Sweet potato mild mottle virus (SPMMV), has a single but large P1 and has HC-Pro. Both virus isolates (TZ:Mlb3:07 & TZ:Kor6:08) characterized in this study contained a single P1 and lacked the HC-Pro which indicates unique evolution in the family Potyviridae. Comparison of 12 complete genomes of CBSD-associated viruses which included two genomes characterized in this study, revealed genetic identity of 69.0–70.3% (nt) and amino acid (aa) identities of 73.6–74.4% at polyprotein level. Comparison was also made among 68 complete CP sequences, which indicated 69.0-70.3 and 73.6-74.4 % identity at nt and aa levels, respectively. The genetic variation was large enough for dermacation of CBSD-associated virus isolates into two distinct species. The name CBSV was retained for isolates that were related to CBSV isolates available in database whereas the new virus described for the first time in this study was named Ugandan cassava brown streak virus (UCBSV) by the International Committee on Virus Taxonomy (ICTV). The isolates TZ:Mlb3:07 and TZ:Kor6:08 belong to UCBSV and CBSV, respectively. The isolates of CBSV and UCBSV were 79.3-95.5% and 86.3-99.3 % identitical at nt level, respectively, suggesting more variation amongst CBSV isolates. The main sources of variation in plant viruses are mutations and recombination. Signals for recombination events were detected in 50% of isolates of each virus. Recombination events were detected in coding and non-coding (3’-UTR) sequences except in the 5’UTR and P3. There was no evidence for recombination between isolates of CBSV and UCBSV. The non-synonomous (dN) to synonomous (dS) nucleotide substitution ratio (ω) for the HAM1h and CP domains of both viruses were ≤ 0.184 suggesting that most sites of these proteins were evolving under strong purifying selection. However, there were individual amino acid sites that were submitted to adaptive evolution. For instance, adaptive evolution was detected in the HAM1h of UCBSV (n=15) where 12 aa sites were under positive selection (P< 0.05) but not in CBSV (n=12). The CP of CBSV (n=23) contained 12 aa sites (p<0.01) while only 5 aa sites in the CP gene of UCBSV were predicted to be submitted to positive selection pressure (p<0.01). The advantages offered by the aa sites under positive selection could not be established but occurrence of such sites in the terminal ends of UCBSV-HAMIh, for example, was interpreted as a requirement for proteolysis during polyprotein processing. Two different primer pairs that simultaneously detect UCBSV and CBSV isolates were developed in this study. They were used successfully to study distribution of CBSV, UCBSV and their mixed infections in Tanzania and Uganda. It was established that the two viruses co-infect cassava and that incidences of co-infection could be as high as 50% around Lake Victoria on the Tanzanian side. Furthermore, it was revealed for the first time that both UCBSV and CBSV were widely distributed in Eastern Africa. The primer pair was also used to confirm infection in a close relative of cassava, Manihot glaziovii (Müller Arg.) with CBSV. DNA barcoding of M. glaziovii was done by sequencing the matK gene. Two out of seven M. glaziovii from the coastal areas of Korogwe and Kibaha in north eastern Tanzania were shown to be infected by CBSV but not UCBSV isolates. Detection in M. glaziovii has an implication in control and management of CBSD as it is likely to serve as virus reservoir. This study has contributed to the understanding of evolution of CBSV and UCBSV, which cause CBSD epidemic in Eastern Africa. The detection tools developed in this work will be useful in plant breeding, verification of the phytosanitary status of materials in regional and international movement of germplasm, and in all diagnostic activities related to management of CBSD. Whereas there are still many issues to be resolved such as the function and biological significance of HAM1h and its origin, this work has laid a foundation upon which the studies on these aspects can be based.