Reassortant Avian Influenza A(H9N2) viruses in chickens in retail poultry shops, Pakistan, 2009-2010.

Phylogenetic analysis of influenza viruses collected during December 2009-February 2010 from chickens in live poultry retail shops in Lahore, Pakistan, showed influenza A(H9N2) lineage polymerase and nonstructural genes generate through inter- and intrasubtypic reassortments. Many amino acid signatures observed were characteristic of human isolates; hence, their circulation could enhance inter- or intrasubtypic reassortment.

The prevalence of brucellosis and bovine tuberculosis in ruminants in Sidi Kacem Province, Morocco.

Bovine tuberculosis (BTB) and brucellosis are major endemic zoonoses in ruminants in Morocco that impact on both animal and human health. This study presents an assessment of the epidemiological and socioeconomic burden of bacterial zoonoses in Sidi Kacem Province in Northern Morocco from a cross-sectional survey of 125 cattle and/or small ruminant-owning households. In total, 1082 sheep and goats were examined from 81 households. The single intradermal comparative cervical test to screen for bovine tuberculosis was undertaken on 1194 cattle from 123 households and all cattle were blood sampled. Cattle and small ruminant sera were tested for brucellosis using the standard Rose Bengal Test (sRBT) and the modified Rose Bengal Test (mRBT). Bacteriology was performed on 21 milk samples obtained from cattle that were seropositive for brucellosis for isolation and phenotyping of circulating Brucella strains. Individual and herd prevalence for BTB in cattle of 20.4% (95% CI 18%-23%) and 57.7% (95% CI 48%-66%), respectively, were observed in this study. The prevalence of brucellosis in cattle at individual and herd level was 1.9% (95% CI 1.2%-2.8%) and 9% (95% CI 4.5%-1.5%), respectively. Brucella pathogens were isolated from three cattle milk samples and were identified as B. abortus using Bruceladder® multiplex PCR and B. abortus biovar 1 by classical phenotyping. All small ruminants were seronegative to sRBT, two were positive to mRBT. A higher risk of BTB and brucellosis was observed in cattle in intensive livestock systems, in imported and crossed breeds and in animals from larger herds (>15). The three risk factors were usually present in the same herds, leading to higher transmission risk and persistence of both zoonoses. These results highlight the importance of implementing control strategies for both BTB and brucellosis to reduce productivity losses and the risk of transmission to humans. Prioritising control for BTB and brucellosis in intensive livestock production systems is essential for human and animal health.

Polisye Kont Moustik: A Culturally Competent Approach to Larval Source Reduction in the Context of Lymphatic Filariasis and Malaria Elimination in Haiti.

Community engagement has become an increasingly important focus of global health programs. Arbovirus emergence in the Americas (Zika and chikungunya virues), and global goals for malaria and lymphatic filariasis elimination, mean that community-based mosquito control has taken on a new salience. But how should mosquito control initiatives be designed and implemented in ways that best engage local people? What are the challenges and trade-offs of different strategies, not only for effectiveness but also for scale-up? In this paper, we describe the social and political dynamics of a pilot study in a small town in northern Haiti. With the aim of developing a culturally-competent approach to larval source management (LSM), our pilot project combined larval surveillance with environmental management, social engagement, community education, and larvicide application. Orientated around a network of 'Mosquito Police' (Polisye Kont Moustik, in Haitian Creole), our approach integrated elements of formative research, social learning, and community participation. Here, we reflect on the challenges we encountered in the field, from larval mapping, staff management, education and behavior change, engagement with formal and informal leaders, and community-based environmental cleanup. We discuss how these programmatic efforts were influenced and shaped by a complex range of social, cultural, political, and economic realities, and conclude by discussing the implications of our community-based approach for the elimination of lymphatic filariasis and malaria, and other vector-borne diseases, in Haiti.

A case-control study to identify risk factors associated with avian influenza subtype H9N2 on commercial poultry farms in Pakistan.

A 1:1 matched case-control study was conducted to identify risk factors for avian influenza subtype H9N2 infection on commercial poultry farms in 16 districts of Punjab, and 1 administrative unit of Pakistan. One hundred and thirty-three laboratory confirmed positive case farms were matched on the date of sample submission with 133 negative control farms. The association between a series of farm-level characteristics and the presence or absence of H9N2 was assessed by univariable analysis. Characteristics associated with H9N2 risk that passed the initial screening were included in a multivariable conditional logistic regression model. Manual and automated approaches were used, which produced similar models. Key risk factors from all approaches included selling of eggs/birds directly to live bird retail stalls, being near case/infected farms, a previous history of infectious bursal disease (IBD) on the farm and having cover on the water storage tanks. The findings of current study are in line with results of many other studies conducted in various countries to identify similar risk factors for AI subtype H9N2 infection. Enhancing protective measures and controlling risks identified in this study could reduce spread of AI subtype H9N2 and other AI viruses between poultry farms in Pakistan.

Candidate protein selection for diagnostic markers of African trypanosomiasis.

The ability to accurately diagnose the presence of an infective micro-organism is not only important for individual human and animal health and wellbeing, but is also central to surveillance programmes. Effective and sustainable control of many diseases in the developing world depends on the availability of field applicable diagnostics that are cheap, reliable, simple in design and application, and which provide immediate results. This review examines how the genome sequences can be used in the selection of potential candidate proteins for developing new serodiagnostics for African trypanosomiasis.

Brucellosis as an emerging threat in developing economies: lessons from Nigeria.

Nigeria is the most populous country in Africa, has a large proportion of the world's poor livestock keepers, and is a hotspot for neglected zoonoses. A review of the 127 accessible publications on brucellosis in Nigeria reveals only scant and fragmented evidence on its spatial and temporal distribution in different epidemiological contexts. The few bacteriological studies conducted demonstrate the existence of Brucella abortus in cattle and sheep, but evidence for B. melitensis in small ruminants is dated and unclear. The bulk of the evidence consists of seroprevalence studies, but test standardization and validation are not always adequately described, and misinterpretations exist with regard to sensitivity and/or specificity and ability to identify the infecting Brucella species. Despite this, early studies suggest that although brucellosis was endemic in extensive nomadic systems, seroprevalence was low, and brucellosis was not perceived as a real burden; recent studies, however, may reflect a changing trend. Concerning human brucellosis, no studies have identified the Brucella species and most reports provide only serological evidence of contact with Brucella in the classical risk groups; some suggest brucellosis misdiagnoses as malaria or other febrile conditions. The investigation of a severe outbreak that occurred in the late 1970s describes the emergence of animal and human disease caused by the settling of previously nomadic populations during the Sahelian drought. There appears to be an increasing risk of re-emergence of brucellosis in sub-Saharan Africa, as a result of the co-existence of pastoralist movements and the increase of intensive management resulting from growing urbanization and food demand. Highly contagious zoonoses like brucellosis pose a threat with far-reaching social and political consequences.

Using molecular data for epidemiological inference: assessing the prevalence of Trypanosoma brucei rhodesiense in tsetse in Serengeti, Tanzania.

BACKGROUND: Measuring the prevalence of transmissible Trypanosoma brucei rhodesiense in tsetse populations is essential for understanding transmission dynamics, assessing human disease risk and monitoring spatio-temporal trends and the impact of control interventions. Although an important epidemiological variable, identifying flies which carry transmissible infections is difficult, with challenges including low prevalence, presence of other trypanosome species in the same fly, and concurrent detection of immature non-transmissible infections. Diagnostic tests to measure the prevalence of T. b. rhodesiense in tsetse are applied and interpreted inconsistently, and discrepancies between studies suggest this value is not consistently estimated even to within an order of magnitude. METHODOLOGY/PRINCIPAL FINDINGS: Three approaches were used to estimate the prevalence of transmissible Trypanosoma brucei s.l. and T. b. rhodesiense in Glossina swynnertoni and G. pallidipes in Serengeti National Park, Tanzania: (i) dissection/microscopy; (ii) PCR on infected tsetse midguts; and (iii) inference from a mathematical model. Using dissection/microscopy the prevalence of transmissible T. brucei s.l. was 0% (95% CI 0-0.085) for G. swynnertoni and 0% (0-0.18) G. pallidipes; using PCR the prevalence of transmissible T. b. rhodesiense was 0.010% (0-0.054) and 0.0089% (0-0.059) respectively, and by model inference 0.0064% and 0.00085% respectively. CONCLUSIONS/SIGNIFICANCE: The zero prevalence result by dissection/microscopy (likely really greater than zero given the results of other approaches) is not unusual by this technique, often ascribed to poor sensitivity. The application of additional techniques confirmed the very low prevalence of T. brucei suggesting the zero prevalence result was attributable to insufficient sample size (despite examination of 6000 tsetse). Given the prohibitively high sample sizes required to obtain meaningful results by dissection/microscopy, PCR-based approaches offer the current best option for assessing trypanosome prevalence in tsetse but inconsistencies in relating PCR results to transmissibility highlight the need for a consensus approach to generate meaningful and comparable data.

No gold standard estimation of the sensitivity and specificity of two molecular diagnostic protocols for Trypanosoma brucei spp. in Western Kenya.

African animal trypanosomiasis is caused by a range of tsetse transmitted protozoan parasites includingTrypanosoma vivax, Trypanosoma congolense and Trypansoma brucei. In Western Kenya and other parts of East Africa two subspecies of T. brucei, T.b. brucei and the zoonoticT.b. rhodesiense, co-circulate in livestock. A range of polymerase chain reactions (PCR) have been developed as important molecular diagnostic tools for epidemiological investigations of T. brucei s.l. in the animal reservoir and of its zoonotic potential. Quantification of the relative performance of different diagnostic PCRs is essential to ensure comparability of studies. This paper describes an evaluation of two diagnostic test systems for T. brucei using a T. brucei s.l. specific PCR [1] and a single nested PCR targeting the Internal Transcribed Spacer (ITS) regions of trypanosome ribosomal DNA [2]. A Bayesian formulation of the Hui-Walter latent class model was employed to estimate their test performance in the absence of a gold standard test for detecting T.brucei s.l. infections in ear-vein blood samples from cattle, pig, sheep and goat populations in Western Kenya, stored on Whatman FTA cards. The results indicate that the system employing the T. brucei s.l. specific PCR (Se1=0.760) had a higher sensitivity than the ITS-PCR (Se2=0.640); both have high specificity (Sp1=0.998; Sp2=0.997). The true prevalences for livestock populations were estimated (pcattle=0.091, ppigs=0.066, pgoats=0.005, psheep=0.006), taking into account the uncertainties in the specificity and sensitivity of the two test systems. Implications of test performance include the required survey sample size; due to its higher sensitivity and specificity, the T. brucei s.l. specific PCR requires a consistently smaller sample size than the ITS-PCR for the detection of T. brucei s.l. However the ITS-PCR is able to simultaneously screen samples for other pathogenic trypanosomes and may thus be, overall, a better choice of test in multi-organism studies.

Patterns in age-seroprevalence consistent with acquired immunity against Trypanosoma brucei in Serengeti lions.

Trypanosomes cause disease in humans and livestock throughout sub-Saharan Africa. Although various species show evidence of clinical tolerance to trypanosomes, until now there has been no evidence of acquired immunity to natural infections. We discovered a distinct peak and decrease in age prevalence of T. brucei s.l. infection in wild African lions that is consistent with being driven by an exposure-dependent increase in cross-immunity following infections with the more genetically diverse species, T. congolense sensu latu. The causative agent of human sleeping sickness, T. brucei rhodesiense, disappears by 6 years of age apparently in response to cross-immunity from other trypanosomes, including the non-pathogenic subspecies, T. brucei brucei. These findings may suggest novel pathways for vaccinations against trypanosomiasis despite the notoriously complex antigenic surface proteins in these parasites.

Factors affecting trypanosome maturation in tsetse flies.

Trypanosoma brucei brucei infections which establish successfully in the tsetse fly midgut may subsequently mature into mammalian infective trypanosomes in the salivary glands. This maturation is not automatic and the control of these events is complex. Utilising direct in vivo feeding experiments, we report maturation of T. b. brucei infections in tsetse is regulated by antioxidants as well as environmental stimuli. Dissection of the maturation process provides opportunities to develop transmission blocking vaccines for trypanosomiasis. The present work suggests L-cysteine and/or nitric oxide are necessary for the differentiation of trypanosome midgut infections in tsetse.

Trypanosoma brucei MOB1 is required for accurate and efficient cytokinesis but not for exit from mitosis.

Two MOB1 genes, MOB1-A and MOB1-B, were identified in Trypanosoma brucei. MOB1-A of T. brucei was shown to form a complex with TbPK50, a functional homologue of the Schizosaccharomyces pombe protein kinase Orb6, and immune precipitated MOB1-A exhibited histone H1 protein kinase activity. MOB1-A and TbPK50 were also shown to bind p12cks1, a cyclin-dependent kinase accessory protein. Immune fluorescence of epitope-tagged MOB1-A and MOB1-B in bloodstream form trypanosomes showed they had a punctate distribution all through the cell cytoplasm and were excluded from the nucleus throughout the cell cycle. Using RNA interference (RNAi), MOB1 was shown to be essential in both bloodstream and procyclic life cycle stages. In the bloodstream form, RNAi of MOB1 resulted, after 8 h, in a significant increase in post-mitotic cells, the majority of which had a visible cleavage furrow. This was followed by the appearance of cells with abnormal complements of nuclei and kinetoplasts, often with the number of nuclei exceeding the number of kinetoplasts. Thus, downregulation of MOB1 in the bloodstream form results in a delay in cytokinesis, and leads to a deregulation of the cell cycle, possibly through an inhibitory effect on kinetoplast replication. In contrast, downregulation of MOB1 in the procyclic form appears to impede the accuracy of cytokinesis, by allowing mispositioning of the cleavage furrow and inappropriate cytokinesis. Unlike its counterpart in budding yeast, T. brucei MOB1 does not appear to be required for mitotic exit.

Theileria annulata: virulence and transmission from single and mixed clone infections in cattle.

Theoretically, parasite virulence should be higher for faster growing parasites, and higher in mixed infections compared to single-clone infections. Virulence should also be positively correlated to transmission rates. Theileria annulata provides a good model system for studying such hypotheses, as parasite replication causes harm to the host, and there is evidence suggesting that the genetic complexity of an infection might affect its virulence. Two clones of T. annulata were chosen, one fast growing and one slow growing in vitro and these were used to establish cattle infections, either alone, or in a mixed infection. Virulence was measured using lymph node expansion, temperature, and blood parameters as correlates. As predicted, the faster growing clone was found to produce higher virulence. Mixed infections did not show higher virulence than single-clone infections, but interactions within mixed infections resulted in more transmission stage production than seen in either of the single-clone infections. Index Descriptors and Abbreviations. Theileria annulata, Apicomplexa, mixed infections, virulence, growth rates, red blood cell, RBC; packed cell volume, PCV.