Targeting the tick/pathogen interface for developing new anaplasmosis vaccine strategies

Bovine anaplasmosis is a tick-borne hemolytic disease of cattle that occurs worldwide caused by the intraerythrocytic rickettsiae Anaplasma marginale. Control measures, including use of acaricides, administration of antibiotics and vaccines, have varied with geographic location. Our research is focused on the tick-pathogen interface for development of new vaccine strategies with the goal of reducing anaplasmosis, tick infestations and the vectorial capacity of ticks. Toward this approach, we have targeted (1) development of an A. marginale cell culture system to provide a non-bovine antigen source, (2) characterization of an A. marginale adhesion protein, and (3) identification of key tick protective antigens for reduction of tick infestations. A cell culture system for propagation of A. marginale was developed and provided a non-bovine source of A. marginale vaccine antigen. The A. marginale adhesion protein, MSP1a, was characterized and use of recombinant MSP1a in vaccine formulations reduced clinical anaplasmosis and infection levels in ticks that acquired infection on immunized cattle. Most recently, we identified a tick-protective antigen, subolesin, that reduced tick infestations, as well as the vectorial capacity of ticks for acquisition and transmission of A marginale. This integrated approach to vaccine development shows promise for developing new strategies for control of bovine anaplasmosis.     

Anaplasmosis in Cattle in Italy

Bovine anaplasmosis caused by Anaplasma marginale is a disease transmitted by ticks belonging to the Ixodidae family. Southern Italy is considered an endemic zone but environmental and social factors are changing the epidemiology of the disease to expand to previously anaplasmosis-free regions. The available data of published reports of anaplasmosis in Italy together with the data obtained by the National Centre of Reference for Anaplasma, Babesia, Rickettsia and Theileria (C.R.A.Ba.R.T.), allowed to report A. marginale infection in different Italian regions (Lazio, Marche, Campania, Puglia, Basilicata, Calabria, Lombardy, Tuscany, Umbria and Sicily). Cattle are also subject to infection with the related Ixodes ricinus-transmitted pathogen, Anaplasma phagocytophilum that results in reduced milk production in cattle. A. phagocytophilum infect also small ruminants, domestic and wild animals and causes the human granulocytic anaplasmosis. Different studies have been conducted on the presence of A. phagocytophilum in Italy both in the tick vectors and in the wild and domestic reservoirs. Contrary to A. marginale, the prevalence of A. phagocytophilum embraces the whole Italian territory from the Alps to the southern and insular regions.     

Tick Vaccines and the Transmission of Tick-Borne Pathogens

Ticks transmit pathogens that cause diseases which greatly impact both human and animal health. Vaccines developed against Boophilus spp. using Bm86 and Bm95 tick gut antigens demonstrated the feasibility of using vaccines for control of tick infestations. These vaccines also reduced transmission of tick-borne pathogens by decreasing exposure of susceptible hosts to ticks. The recently discovered tick antigens, 64P putative cement protein and subolesin involved in the regulation of tick feeding and reproduction, were also shown to reduce tick infestations. These antigens, together with the TROSPA receptor for Burrelia burgdorferi OspA were effective against tick-borne pathogens by reducing the infection levels in ticks and/or the transmission of the pathogen. Development of a vaccine targeted at both the tick vector and pathogen would contribute greatly to the control of tick infestations and the transmission of tick-borne diseases. These results have demonstrated that tick vaccines can be developed for control tick infestations and show promise for the prevention of the transmission of tick-borne pathogens.     

Prevalence and risk factors associated with <Emphasis Type="Italic">Theileria parva</Emphasis> infection in cattle in three regions of Tanzania

Ticks and tickborne diseases (TBDs) are serious constraints to cattle production in Tanzania and other tropical and subtropical countries. Among the TBDs, East Coast fever (ECF) is the most important as it causes significant economic losses to the cattle industry in Tanzania. However, control of ECF in Tanzania has continued to be a challenge due to inadequate epidemiological information. The main objective of this study was to determine the epidemiological situation of Theileria parva infections in cattle kept under pastoral and agro-pastoral farming systems in Mara, Singida, and Mbeya regions of Tanzania. Blood samples were collected from 648 cattle in the three regions. Genomic DNA was extracted and amplified in a polymerase chain reaction (PCR) using T. parva-specific primers targeting the 104-kD antigen (P104) gene. In addition, information was collected on the possible risk factors of T. parva infection (animal age, region, animal sex, tick burden, tick control method, and frequency of acaricide application). The prevalence of T. parva across the three regions was 14.2%. There was variation in prevalence among the three regions with Mara (21.8%) having a significantly higher (p = 0.001) prevalence than the other regions. Moreover, Mbeya exhibited relatively lower prevalence (7.4%) compared to the other regions. Factors found to be significantly associated with an animal being PCR positive for T. parva were region (p = 0.001) and tick burden (p = 0.003). Other factors were not found to be significant predictors of being PCR positive for T. parva. The present study showed high variation in tick burden and T. parva prevalence across the regions. Therefore, different strategic planning and cost-effective control measures for ticks and T. parva infection should be implemented region by region in order to reduce losses caused by ticks and ECF in the study area.     

An <Emphasis Type="Italic">in-vivo</Emphasis> study of the efficacy and safety of ethno-veterinary remedies used to control cattle ticks by rural farmers in the Eastern Cape Province of South Africa

Ticks feed on blood, are vectors of tick-borne diseases and cause considerable skin damage to livestock. They are commonly controlled using commercial acaricides, which are expensive to the rural farmers, causing them to resort to alternative tick control methods. The objective of this study was to assess the acaricidal properties and safety of some materials (Ptaeroxylon obliquum, Aloe ferox, Lantana camara, Tagetes minuta, Used engine oil and Jeyes fluid, used by rural farmers to control cattle ticks in the Eastern Cape Province, South Africa. A total of 52 cattle were divided into 13 experimental groups with 4 cattle in each. Jeyes fluid at 76.8% concentration and Used engine oil had an efficacy that was almost similar to that of the positive control Ektoban® (Cymiazol 17.5 and cypermethrin 2.5%). Extracts of L. camara at 40% concentration had an efficacy of 57% while A. ferox, P. obliquum and T. minuta were not effective. The test materials had no irritation effect on rats. The study revealed that the materials rural farmers use as acaricides vary in their efficacy in controlling ticks.     

Seroprevalence of <Emphasis Type="Italic">Anaplasma marginale</Emphasis> and <Emphasis Type="Italic">Babesia bigemina</Emphasis> infections and associated risk factors in Machakos County,Kenya

Anaplasma marginale and Babesia bigemina are important tick-borne pathogens of cattle. A cross-sectional survey was undertaken to determine the seroprevalence of A. marginale and B. bigemina infections and identify associated risk factors on traditional smallholder farms in Machakos County, Kenya. A total of 421 cattle from 127 farms from four divisions in the county were sampled and visited between September and November 2007. The farms were selected by a proportional allocation approach based on the number of farms in the four divisions previously selected by stratified random sampling method. Information on animal and individual farm management variables was obtained using standardized questionnaires. Prevalence of serum antibodies due to A. marginale and B. bigemina pathogens was determined using the enzyme-linked immunosorbent assay (ELISA) technique. The relationship between the seropositivity and associated risk factors was assessed by multivariable analyses using standard logistic regression models. The overall estimation (and their 95% confidence intervals) of A. marginale and B. bigemina seropositivity at the animal level was 53.4% (48.5%, 58.2%) and 40.6% (35.8%, 45.4%), respectively. Two variables, “animal age” and “administrative division,” were significantly associated with the A. marginale seroresponse. Three variables, “animal age” “grazing system” and “administrative division” were significantly associated with the B. bigemina seroresponse. These findings suggest possible indicators of existence of endemic instability for the two infections. The study identifies characterization of environmental suitability for the vectors and how they interact with grazing systems to cause the infections as an area for further studies, for improved understanding of the infections and in designing disease control programs.     

The natural history of Anaplasma marginale

The intracellular pathogen Anaplasma marginale (Rickettsiales: Anaplasmataceae), described by Sir Arnold Theiler in 1910, is endemic worldwide in tropical and subtropical areas. Infection of cattle with A. marginale causes bovine anaplasmosis, a mild to severe hemolytic disease that results in considerable economic loss to both dairy and beef industries. Transmission of A. marginale to cattle occurs biologically by ticks and mechanically by biting flies and by blood-contaminated fomites. Both male ticks and cattle hosts become persistently infected with A. marginale and serve as reservoirs of infection. While erythrocytes are the major site of infection in cattle, A. marginale undergoes a complex developmental cycle in ticks that begins by infection of gut cells, and transmission to susceptible hosts occurs from salivary glands during feeding. Major surface proteins (MSPs) play a crucial role in the interaction of A. marginale with host cells, and include adhesion proteins and MSPs from multigene families that undergo antigenic change and selection in cattle, thus contributing to maintenance of persistent infections. Many geographic strains of A. marginale have been identified worldwide, which vary in genotype, antigenic composition, morphology and infectivity for ticks. Isolates of A. marginale may be maintained by independent transmission events and a mechanism of infection/exclusion in cattle and ticks. The increasing numbers of A. marginale genotypes identified in some geographic regions most likely resulted from intensive cattle movement. However, concurrent A. marginale strain infections in cattle was reported, but these strains were more distantly related. Phylogenetic studies of selected geographic isolates of A. marginale, using msp4 and msp1α, provided information about the biogeography and evolution of A. marginale, and msp1α genotypes appear to have evolved under positive selection pressure. Live and killed vaccines have been used for control of anaplasmosis and both types of vaccines have advantages and disadvantages. Vaccines have effectively prevented clinical anaplasmosis in cattle but have failed to block A. marginale infection. Vaccines are needed that can prevent clinical disease and, simultaneously, prevent infection in cattle and ticks, thus eliminating these hosts as reservoirs of infection. Advances in genomics, proteomics, immunology and biochemical and molecular technologies during the last decade have been applied to research on A. marginale and related organisms, and the recent development of a cell culture system for A. marginale has provided a format for studying the pathogen/tick interface. Recent advancements and new research methodologies should provide additional opportunities for development of new strategies for control and prevention of bovine anaplasmosis.     

Genetic parameters for tick counts across months for different tick species and anatomical locations in South African Nguni cattle

The objective of the study was to characterise genetic parameters across months for different tick species and anatomical locations in South African Nguni cattle. Tick counts were conducted monthly, over a 2-year period, on 586 Nguni cattle under natural infestation, from four herds located in different provinces of South Africa. The counts were recorded for six species of ticks (Amblyomma hebraeum, Rhipicephalus evertsi evertsi, Rhipicephalus decoleratus and microplus (Boofilids), Rhipicephalus appendiculatus, Rhipicephalus simus and Hyalomma marginatum) attached on eight anatomical locations on the animals and were summed by species and anatomical location. Heritability estimates, phenotypic and genetic correlations were estimated on a monthly basis using mixed linear models, fitting univariate and bivariate sire models. Fixed effects considered were location, sex, year and age as a covariate. Tick counts were higher in the hot months, and A. hebraeum was the most dominant tick species. Heritability estimates for tick count varied by month and trait and ranged from 0 to 0.89. Genetic correlations were mostly positive, and low to high, with some negative correlations with high standard error. Phenotypic correlations were low to moderate. In general, high genetic correlations were observed between whole body count and the anatomical location counts, suggesting that it may not be necessary to conduct whole body counts. Counts from the belly and perineum appeared to be the most suitable surrogate traits for whole body count. These findings provide useful information for developing strategies for the practical implementation of genetic selection, as a supplement to the traditional tick control measures.     

Risk factors for anaplasmosis in dairy cows during the peripartum

Anaplasma marginale is endemic in tropical and subtropical areas around the world. Some studies have suggested that cows during peripartum may present a transient immunosuppression state and development of clinical signs of anaplasmosis. The aim of this study was to investigate the relationship between some risk factors and the seroprevalence of A. marginale in dairy cows during peripartum in Rio de Janeiro, Brazil. The risk factors analyzed in association with the prevalence of antibodies against A. marginale in dairy cows were calving season, reproductive experience, breed standard, tick infestations, stocking density, and milk yield. The antibodies against A. marginale were tested in indirect enzyme-linked immunosorbent assays. A primary screening using a 2 × k contingency table of the exposed variables with the outcomes was performed. All variables for which p?<?0.20 were included in a fixed effects log regression. The risk factors investigated to anaplasmosis were calving (OR 2.61, IC 1.08–7.63), breed standard (OR 3.83, IC 0.08–0.28), reproductive experience (OR 33.7, IC 2.14–5.16), milk yield (OR 3.9, IC 2.24–7.03), Rhipicephalus microplus infestations (OR 10.3, IC 0.05–0.17), and stocking density (OR 22.3, IC 0.05–0.17). Low titers of antibodies against A. marginale during peripartum had been characterized as a period previous to development of clinical anaplasmosis. Thus, studies on anaplasmosis should consider each farm as an epidemiological unit, where environmental and immunological factors may influence the endemic status of the pathogen.     

Isolation and characterization of two lytic bacteriophages against <Emphasis Type="Italic">Staphylococcus aureus</Emphasis> from India: newer therapeutic agents against Bovine mastitis

Bovine mastitis causes severe economic losses to dairy farmers. Staphylococcus aureus, is one of the most important pathogen implicated in etiology of clinical and subclinical mastitis in bovines. In view of increasing antimicrobial resistance alternatives to antibiotic therapy are much needed. The present decade has witnessed a renewed interest in phage based therapeutics and diagnostics. The present study, describes isolation and characterization of two lytic phages SAJK-IND and MSP against Staphylococcus aureus having a potential to be used in therapy against mastitis. SAJK-IND and MSP phages belonged to Myoviridae and Podoviridae families, respectively. TEM imaging of the two phages revealed an iscosahedral head. MSP phage has a short non contractile tail. SAJK-IND and MSP have a burst size of 44?±?3 and 25?±?5 PFU/ infected cell, respectively. SAJK-IND and MSP phages revealed ? 12 and ?16 proteins, respectively on SDS-PAGE analysis. The lytic activity of the phages was specific for Staphylococcus aureus. SAJK-IND revealed 100% lytic activity against several strains of Staphylococcus aureus isolated from mastitis milk samples whereas, MSP had only 40% lytic activity. SAJK-IND phage genome was sequenced, assembled and deposited in Genbank under accession no MG010123.     

Use of serology and real time PCR to control an outbreak of bovine brucellosis at a dairy cattle farm in the Nile Delta region,Egypt

Background

Bovine brucellosis remains one of the most prevalent zoonotic infections affecting dairy cattle in developing countries where the applied control programs often fail. We analyzed the epidemiologic pattern of bovine brucellosis in a dairy cattle herd that showed several cases of abortions after regular vaccination with RB51 (B. abortus vaccine). In 2013 thirty dairy cows, from a Holstein-Friesian dairy herd with a population of 600 cattle, aborted five months post vaccination by a regular RB51 vaccine. Blood samples were drawn from milking cows and growing heifers, as well as heifers and cows pregnant up to 6 months. These samples were collected in June 2013 (n?=?257) and May 2014 (n?=?263) and were tested by real time (rt)-PCR as well as serological tests, in particular Rose Bengal Test (RBT), Enzyme-Linked Immunosorbent Assays (ELISA) and Fluorescence Polarization Assay. Tissue specimens were also collected from an aborted fetus and cultured. Isolates were subjected to bacteriological typing tests at the genus and species levels.

Results

Five months post vaccination with RB51 vaccine, Brucella (B.) DNA was detected in blood samples of cows by rt-PCR. The serological tests also revealed the spread of Brucella field strains within the herd in 2013. Four Brucella isolates were recovered from specimens collected from the aborted fetus. These isolates were typed as follows: one B. abortus RB51 vaccine strain and three isolates of B. abortus field strain. The seropositive cows with positive rt-PCR might indicate an infection by the Brucella field strain; while the positive rt-PCR results from seronegative animals may either be due to circulating RB51 vaccine DNA in vaccinated animals or to circulating field strain in infected animals before seroconversion.

Conclusion

The results herein suggest that PCR can be a good supplementary tool in an outbreak situation, if an assay is available that can differentiate vaccine and field strains with a high analytical sensitivity. We recommend using RBT and ELISA in parallel in outbreak situations, to identify as many infected animals as possible during the initial screenings. This test procedure should be repeated for at least three successive negative tests, with one month interval.

     

Transformation of Anaplasma marginale

The tick-borne pathogen, Anaplasma marginale, has a complex life cycle involving ruminants and ixodid ticks. It causes bovine anaplasmosis, a disease with significant economic impact on cattle farming worldwide. The obligate intracellular growth requirement of the bacteria poses a challenging obstacle to their genetic manipulation, a problem shared with other prokaryotes in the genera Anaplasma, Ehrlichia, and Rickettsia. Following our successful transformation of the human anaplasmosis agent, A. phagocytophilum, we produced plasmid constructs (a transposon bearing plasmid, pHimarAm-trTurboGFP-SS, and a transposase expression plasmid, pET28Am-trA7) designed to mediate random insertion of the TurboGFP and spectinomycin/streptomycin resistance genes by the Himar1 allele A7 into the A. marginale chromosome. In these trans constructs, expression of the fluorescent and the selectable markers on the transposon, and expression of the transposase are under control of the A. marginale tr promoter. Constructs were co-electroporated into A. marginale St. Maries purified from tick cell culture, and bacteria incubated for 2 months under selection with a combination of spectinomycin and streptomycin. At that time, ≤1% of tick cells contained colonies of brightly fluorescent Anaplasma, which eventually increased to infect about 80–90% of the cells. Cloning of the insertion site in E. coli and DNA sequence analyses demonstrated insertion of the entire plasmid pHimarAm-trTurboGFP-SS encoding the transposon in frame into the native tr region of A. marginale in an apparent single homologous crossover event not mediated by the transposase. Transformants are fastidious and require longer subculture intervals than wild type A. marginale. This result suggests that A. marginale, as well as possibly other species of Anaplasma and Ehrlichia, can be transformed using a strategy of homologous recombination.     

Cryptosporidium species detected in calves and cattle in Dagoretti,Nairobi, Kenya

A total of 1,734 cattle faecal samples from 296 dairy-keeping households were collected from urban settings in Nairobi, Kenya. Modified Ziehl–Neelsen staining method and an immunofluorescence assay were used to identify those samples with Cryptosporidium oocyst infection. Oocysts from positive faecal samples were isolated by Sheather's sucrose flotation method and picked from the concentrate using cover slips. Genomic DNA was extracted from 124 of the faecal samples that were positive for Cryptosporidium and was used as template for nested PCR of the 18S rRNA gene. Twenty-five samples (20 %) were PCR-positive for Cryptosporidium, and 24 of the PCR products were successfully cloned and sequenced. Sequence and phylogenetic analysis identified 17 samples (68 %) as Cryptosporidium parvum-like, four samples (16 %) as Cryptosporidium ryanae, three samples (12 %) as Cryptosporidium andersoni and one sample (4 %) as Cryptosporidium hominis. To the best of our knowledge, this is the first genotyping study to report C. parvum-like, C. andersoni and C. hominis in cattle from Kenya. The results of this study show Cryptosporidium infections in calves and cattle may be potential zoonotic reservoirs of the parasite that infects humans.     

The use of tick transmission by Boophilus microplus to isolate pure strains of Babesia bovis,Babesia bigemina and Anaplasma marginale from cattle with mixed infections

Pure strains of Babesia bovi, Babesia bigemina and Anaplasma marginale were isolated from cattle infected with all 3 species as well as a Theileria sp. and Eperythrozoon teganodes, using only transmission by the tick, Boophilus microplus. Unengorged adult ticks transferred to susceptible cattle transmitted A. marginale, but not Babesia. Engorged adults gave rise to progeny that transmitted Babesia, B. bovis by larvae and B. bigemina by male ticks. The Theileria and E. teganodes were not transmitted by the ticks and thus did not appear in calves used for isolating the pure strains of Babesia and A. marginale.     

Antimicrobial resistance and virulence profile of enterococci isolated from poultry and cattle sources in Nigeria

This study investigated the occurrence, antimicrobial resistance and virulence of Enterococcus from poultry and cattle farms. Three hundred and ninety samples: cloacal/rectal swabs (n?=?260) and manure (n?=?130] were processed for recovery of Enterococcus species. Standard bacteriological methods were used to isolate, identify and characterize Enterococcus species for antimicrobial susceptibility and expression of virulence traits. Detection of antibiotic resistance and virulence genes was carried out by polymerase chain reaction. Enterococcus was recovered from 167 (42.8%) of the 390 samples tested with a predominance of Enterococcus faecium (27.7%). Other species detected were Enterococcus gallinarum, Enterococcus faecalis, Enterococcus hirae, Enterococcus raffinosus, Enterococcus avium, Enterococcus casseliflavus, Enterococcus mundtii and Enterococcus durans. All the isolates tested were susceptible to vancomycin, but resistance to tetracycline, erythromycin, ampicillin and gentamicin was also observed among 61.0, 61.0, 45.1 and 32.7% of the isolates, respectively. Sixty (53.1%) of the isolates were multidrug resistant presenting as 24 different resistance patterns with resistance to gentamicin-erythromycin-streptomycin-tetracycline (CN-ERY-STR-TET) being the most common (n?=?11) pattern. In addition to expression of virulence traits (haemolysin, gelatinase, biofilm production), antibiotic resistance (tetK, tetL, tetM, tetO and ermB) and virulence (asa1, gelE, cylA) genes were detected among the isolates. Also, in vitro transfer of resistance determinants was observed among 75% of the isolates tested. Our data revealed poultry, cattle and manure in this area are hosts to varying Enterococcus species harbouring virulence and resistance determinants that can be transferred to other organisms and also are important for causing nosocomial infection.     

Comparison of rapid diagnostic tests to detect <Emphasis Type="Italic">Mycobacterium avium</Emphasis> subsp. <Emphasis Type="Italic">paratuberculosis</Emphasis> disseminated infection in bovine liver

Mycobacterium avium subsp. paratuberculosis (MAP) causes Johne’s disease, a chronic enteritis in cattle and other domestic and wild ruminants. The presence of MAP in tissues other than intestines and associated lymph nodes, such as meat and liver, is a potential public health concern. In the present study, the relationship between the results of rapid diagnostic tests of the Johne’s disease, such as serum ELISA, rectal scraping PCR, and acid-fast staining, and the presence of MAP in liver was evaluated. Blood, liver, and rectal scraping samples were collected from 200 slaughtered cattle with unknown Johne’s disease status. ELISA was performed to determine the MAP antibody activity in the serum. Acid-fast staining was performed on rectal scraping samples, and PCR was performed on rectal scraping and liver samples. PCR-positive liver samples were used for mycobacterial culture. Overall, the results of this study demonstrated that MAP can be detected and cultured from liver of slaughtered cattle and rapid diagnostic tests of Johne’s disease have limited value in detecting cattle with MAP infection in liver. These findings show that the presence of MAP in liver tissue may occur in cows with negative results for rapid diagnostic tests and vice versa. Hence, liver might represent another possible risk of human exposure to MAP. Given concerns about a potential zoonotic role for MAP, these results show the necessity to find new methods for detecting cattle with MAP disseminated infection.     

Immunogenicity of adenovirus and DNA vaccines co-expressing P39 and lumazine synthase proteins of <Emphasis Type="Italic">Brucella abortus</Emphasis> in BALB/c mice

Brucella poses a great threat to animal and human health. Vaccination is the most promising strategy in the effort to control Brucella abortus (B. abortus) infection, but the currently used live vaccines interfere with diagnostic tests and could potentially result in disease outbreak. Therefore, new subunit vaccines and combined immunization strategies are currently under investigation. In this study, immunogenicity and protection ability of a recombinant adenovirus and plasmid DNA vaccine co-expressing P39 and lumazine synthase proteins of B. abortus were evaluated based on the construction of the two molecular vaccines. Four immunization strategies (single adenovirus, single DNA, adenovirus/DNA, DNA/adenovirus) were investigated. The results showed that the immunization strategy of DNA priming followed by adenovirus boosting induced robust humoral and cellular immune responses, and it significantly reduced the numbers of B. abortus in a mouse model. These results suggest that it could be a potential antigen candidate for development of a new subunit vaccine against B. abortus infection.     

Meat retail conditions within the establishments of Kigali city (Rwanda): bacteriological quality and risk factors for <Emphasis Type="Italic">Salmonella</Emphasis> occurrence

Meat constitutes one of the major vehicles for human foodborne infections. This study aimed to assess the retail conditions and to determine the microbiological quality and safety of meat retailed within the establishments of Kigali (Rwanda). A questionnaire survey was carried out in 150 retail outlets to characterise meat retail conditions. Additionally, 270 retail meat samples were analysed for the enumeration of hygiene indicator bacteria (total mesophilic bacteria and Escherichia coli) and for the qualitative detection of Salmonella, using conventional culture methods. The results revealed that beef was the predominant meat sold within the retail premises of Kigali city, while meat from non-bovine animal species was mainly sold in large establishments. Salmonella was detected in 19.6% of all the retailed meat samples evaluated, whereas the mean loads for total mesophilic bacteria and E. coli were 7.3 and 3.5 log cfu/g, respectively. Three factors, namely the temperature conditions of the meat under retail, the cleanability of the used meat cutting boards, and the training of personnel in hygienic meat handling practices, were found to be significantly (p ≤ 0.05) associated with the risk of Salmonella occurrence in the retailed meat. The findings from this study highlight the need for improvements in hygienic meat handling practices, particularly, in small and medium meat retail establishments in Kigali.     

Prevalence and antimicrobial resistance profiles of <Emphasis Type="Italic">Salmonella enterica</Emphasis> serovars isolated from slaughtered cattle in Bahir Dar,Ethiopia

A study was undertaken from October 2006 to March 2007 to determine the prevalence and antimicrobial resistance patterns of Salmonella serovars. Liver, mesenteric lymph nodes, intestinal content, and carcass swab samples (each n?=?186) were collected from 186 apparently healthy slaughtered cattle at Bahir Dar abattoir. Bacteriological analysis was done according to the International Organization for Standardization (ISO 6579 2002). Isolates were serotyped at Agence Française de Securite Sanitaire des Aliments, Cedex, France. Twenty-eight isolates consisting of Salmonella Typhimurium, Salmonella Newport, Salmonella Haifa, Salmonella Heidelberg, Salmonella Infantis, and Salmonella Mishmarhaemek were identified. Salmonella Typhimurium and Salmonella Newport were most frequently isolated while Salmonella Heidelberg and Salmonella Mishmarhaemek were isolated least. Eleven of the 28 (39.3%) were resistant to one or more of the antimicrobials tested. Resistance was shown to ampicillin, chloramphenicol, gentamycin, norfloxacin, polymyxin-B, streptomycin, tetracycline, and trimethoprim. Four of 11 (36.4%) were multiple antimicrobial resistant. All the isolates tested were susceptible to the antimicrobial effects of gentamycin, norfloxacin, and trimethoprim. Eleven, four, and two isolates of the 28 were resistant to streptomycin, tetracycline, and ampicillin, respectively. All isolates of Salmonella Infantis, Salmonella Typhimurium (except one), and Salmonella Mishmarhaemek were susceptible to the tested antimicrobials. One Typhimurium isolate was resistant to chloramphenicol, streptomycin, and tetracycline. Salmonella Haifa was multiply antimicrobial resistant to ampicillin, tetracycline, and streptomycin. All isolates of Salmonella Heidelberg were resistant to streptomycin. Results of this study indicated high level of carcass contamination with antimicrobial-resistant Salmonella serovars which could pose public health risk; suggests need for hygienic slaughtering operations and proper cooking of meat before consumption. Further detailed studies involving different abattoirs, animal products, food items, and animals on different settings were recommended in the study area.     

Improving smallholder cattle reproductive efficiency in Cambodia to address expanding regional beef demand

This study aimed to identify factors associated with cattle reproductive output in rural smallholder farms in Cambodia in order to determine the main causes of reproductive failure and design efficient interventions for improvement. The majority of the nation’s beef is produced on smallholder farms where productivity is constrained by poor animal reproductivity reflected in the recent livestock population decline of approximately 13 % from 2009 to 2013. Farmers (n?=?240) from 16 villages from five provinces were surveyed in mid-2015 to determine their baseline knowledge, attitude and practices (KAP) associated with cattle reproduction. In addition, 16 case studies from three of these provinces were conducted to provide a more detailed assessment of current cattle reproductive husbandry practices. In order to assess the reproductive impact of previously implemented interventions, an endpoint KAP survey and longitudinal health and husbandry study from three Cambodian provinces conducted between 2008 and 2013 were also analysed. Three multivariable prediction models (two KAP and one longitudinal) identified the following significant factors associated with the reproductive outcomes ‘number of calves born’ or probability that cows ‘gave birth’: target feeding (P?=?0.074), growing vegetables (P?=?0.005), attitudes towards cattle vaccination (P?=?0.010), improving bull selection (P?=?0.032), local breed use (P?=?0.005), number of joining attempts (P?<?0.001), discontinuation of animal draught practices (P?=?0.003) and retention of breeding animals (P?<?0.001). The identification of significant factors and interventions in this study has led to intervention recommendations that can potentially improve reproductive efficiency, combat the declining cattle population and improve smallholder capacity to supply to expanding regional meat demand in South-East Asia and China.