Reservoir For Human Infection Research Articles

Investigation of On-Farm Transmission Routes for Contamination of Dairy Cows with Top 7 Escherichia coli O-Serogroups.

Shiga toxin-producing Escherichia coli (STEC) are foodborne bacterial pathogens, with cattle a significant reservoir for human infection. This study evaluated environmental reservoirs, intermediate hosts and key pathways that could drive the presence of Top 7 STEC (O157:H7, O26, O45, O103, O111, O121 and O145) on pasture-based dairy herds, using molecular and culture-based methods. A total of 235 composite environmental samples (including soil, bedding, pasture, stock drinking water, bird droppings and flies and faecal samples of dairy animals) were collected from two dairy farms, with four sampling events on each farm. Molecular detection revealed O26, O45, O103 and O121 as the most common O-serogroups, with the greatest occurrence in dairy animal faeces (> 91%), environments freshly contaminated with faeces (> 73%) and birds and flies (> 71%). STEC (79 isolates) were a minor population within the target O-serogroups in all sample types but were widespread in the farm environment in the summer samplings. Phylogenetic analysis of whole genome sequence data targeting single nucleotide polymorphisms revealed the presence of several clonal strains on a farm; a single STEC clonal strain could be found in several sample types concurrently, indicating the existence of more than one possible route for transmission to dairy animals and a high rate of transmission of STEC between dairy animals and wildlife. Overall, the findings improved the understanding of the ecology of the Top 7 STEC in open farm environments, which is required to develop on-farm intervention strategies controlling these zoonoses.

Giardia and Cryptosporidium infections in Danish cats: risk factors and zoonotic potential.

Giardia and Cryptosporidium infections are common in cats, but knowledge is limited about their clinical importance, risk factors, and the role of cats as a reservoir for human infections. Here, we collected faeces and questionnaire data from 284 cats from shelters and veterinary clinics in the Copenhagen Metropolitan Region (= study population). Additionally, 33 samples were analysed separately from catteries with gastrointestinal clinical signs (= cases). (Oo-)cysts were quantified by immunofluorescence microscopy. All Giardia (n= 34) and Cryptosporidium (n= 29) positive samples were analysed by sequencing of the 18S rRNA, gdh and hsp70 loci, and co-infections were detected by McMaster/inverted microscopy. In the study population, 7.0% and 6.7% were positive for Giardia and Cryptosporidium respectively; 48.5% and 36.4% of the breeder cats (cases) were infected. Increased odds of diarrhoea were demonstrated in Giardia (p= 0.0008) and Cryptosporidium (p= 0.034) positive cats. For Giardia, the odds were positively correlated with infection intensity. Co-infection with Cryptosporidium (OR 12.79; p< 0.001), parasitic co-infections other than Cryptosporidium (OR 5.22; p= 0.009), no deworming (OR 4.67; p= 0.035), and male sex (OR 3.63; p= 0.025) were risk factors for Giardia. For Cryptosporidium, co-infection with Giardia was the only risk factor (OR 11.93; p< 0.0001). Genotyping revealed G. duodenalis assemblages A and F, and C. felis, all of them previously detected in humans. In conclusion, excretion of Giardia and Cryptosporidium was associated with clinical disease. Although a public health risk is likely, studies including larger sample sizes, more discriminatory markers and samples from other animals and humans are needed to reveal the full zoonotic potential.

Increased risk of hepatitis E virus infection in workers occupationally exposed to sheep.

Hepatitis E virus (HEV) is an enteric RNA virus from the family Hepeviridae with five genotypes (genotypes 1-4 and 7) known to infect humans. HEV infection is known to have a zoonotic swine origin in industrialized countries. The role of pigs and wild boars as major reservoirs for human infection is today well-established; however, the list of new animal reservoirs is ever-expanding as new HEV strains are continuously being found in a broad host range. The recent detection of HEV in sheep stools brings concerns on the possibility of HEV transmission from these animals to humans, particularly in those occupationally exposed. The present work investigated the potential occupational risk of HEV infection in shepherds and sheep milk cheesemakers-workers occupationally exposed to ovine (WOEOs; N=96)-from a region of the Centre of Portugal ('Serra da Estrela') based on the differences of anti-HEV IgG seroprevalence rates between these professionals and the general population (N=192). The presence of HEV-specific antibodies in sheep (N=90) from the same region was also evaluated. The HEV seroprevalence in WOEOs (29.3%) was found to be significantly higher (p=.0198) when compared with population controls (16.1%) which suggests an increased risk for HEV infection in these workers. HEV-specific antibodies were also found in 16.6% of the studied sheep showing that HEV circulates in these animals. Further studies are needed to confirm the zoonotic potential of sheep HEV.

Genomic analyses of Burkholderia cenocepacia reveal multiple species with differential host-adaptation to plants and humans

BackgroundBurkholderia cenocepacia is a human opportunistic pathogen causing devastating symptoms in patients suffering from immunodeficiency and cystic fibrosis. Out of the 303 B. cenocepacia strains with available genomes, the large majority were isolated from a clinical context. However, several isolates originate from other environmental sources ranging from aerosols to plant endosphere. Plants can represent reservoirs for human infections as some pathogens can survive and sometimes proliferate in the rhizosphere. We therefore investigated if B. cenocepacia had the same potential.ResultsWe selected genome sequences from 31 different strains, representative of the diversity of ecological niches of B. cenocepacia, and conducted comparative genomic analyses in the aim of finding specific niche or host-related genetic determinants. Phylogenetic analyses and whole genome average nucleotide identity suggest that strains, registered as B. cenocepacia, belong to at least two different species. Core-genome analyses show that the clade enriched in environmental isolates lacks multiple key virulence factors, which are conserved in the sister clade where most clinical isolates fall, including the highly virulent ET12 lineage. Similarly, several plant associated genes display an opposite distribution between the two clades. Finally, we suggest that B. cenocepacia underwent a host jump from plants/environment to animals, as supported by the phylogenetic analysis. We eventually propose a name for the new species that lacks several genetic traits involved in human virulence.ConclusionRegardless of the method used, our studies resulted in a disunited perspective of the B. cenocepacia species. Strains currently affiliated to this taxon belong to at least two distinct species, one having lost several determining animal virulence factors.

Seasonal and spatial variation in Toxoplasma gondii contamination in soil in urban public spaces in California, United States.

Toxoplasma gondii is a zoonotic parasite that can have severe implications for human health. Acutely infected cats shed environmentally resistant T. gondii oocysts in their faeces that contaminate soil, and soil can serve as a reservoir of infection for humans. Free-roaming domestic cats are thought to play an important role in environmental contamination with T. gondii, but few studies have directly measured the direct contribution of free-roaming cats to T. gondii in soil. Our goals were to determine whether T. gondii soil contamination occurs in public areas with free-roaming cat colonies in central California and examine spatial and temporal variation in soil contamination. We initially performed spiking experiments to compare the limit of T. gondii detection in soil using three conventional nested PCR assays and one real-time quantitative PCR. The nested PCR targeting the internal transcribed spacer (ITS-1) of the small subunit ribosomal RNA was the most sensitive assay, with a limit of detection between 20 and 200 oocysts per gram of soil. We applied the ITS1 PCR assay on soil from sites in city and state parks, public playgrounds and community gardens in central California, USA. Samples were collected during spring, summer and fall and in sites located along the coast and inland. We detected and sequence-confirmed T. gondii in 5.6% ofall of our soil sub-samples, but with large seasonal and spatial variation in soil contamination: we only detected T. gondii during fall and only in coastal sites (44.3% soil prevalence), despite similar sampling intensity across space and time. Our results suggest that free-roaming cat colonies are an important source of T. gondii in spaces where people recreate and grow food and that soil contamination is highly seasonal and spatially variable. Management of free-roaming cats could prevent T. gondii infections by reducing environmental contamination with this zoonotic pathogen.

Detection of virulent thermophilic Campylobacter species in communal chickens

Campylobacter is a foodborne pathogen found in the gut of poultry and other animals, which thereby act as reservoirs for human infection. Campylobacter produces various virulence factors which contribute to the establishment of disease. The aim of this study was to detect virulence genes in Campylobacter isolated from chicken faeces. A total of 408 poultry cloacal samples were analysed through culture, PCR and DNA sequencing. Campylobacter were detected in 24 (5.9 %) of the samples, of which 21 were C. jejuni while 3 were C. coli. The cdtB gene was detected in all Campylobacter isolates while cdtA and cdtC genes were detected in 20 (83.3%) and 18 (75.0%) of the isolates, respectively. The cadF gene was detected in 18 (85.7%) C. jejuni but not in C. coli. Eight genotypes designated G1 to G8 were detected among the isolates. Genotypes G1 and G7 possessed all three cdt genes, a potential for producing functionally active toxins. A large proportion (70.8%) of the isolates possessed both genes for toxin production and adhesion. The high prevalence of cdt and cadF genes in Campylobacter species in this study indicates the pathogenic potential of the isolates. There is a need to implement control measures to curb the spread of these virulent strains.&#x0D; Significance:&#x0D; &#x0D; Virulent thermophilic campylobacters cause severe health complications in humans.&#x0D; Although chicken meat is the main source of human infections, there is limited data on the occurrence of Campylobacter in chicken faeces in southern Africa.&#x0D; This study adds to literature on the occurrence of virulent strains of Campylobacter species, notably through the deposition of the DNA sequences of our local strains into GenBank®.&#x0D; Communal chickens may serve as a source of virulent thermophilic campylobacters to humans, resulting in serious public health implications.&#x0D; &#x0D; Open data sets: &#x0D; MF872608 - https://www.ncbi.nlm.nih.gov/nuccore/MF872608.1 MF872609 - https://www.ncbi.nlm.nih.gov/nuccore/MF872609.1MF872610 - https://www.ncbi.nlm.nih.gov/nuccore/MF872610.1MF872611 - https://www.ncbi.nlm.nih.gov/nuccore/MF872611.1MF872612 - https://www.ncbi.nlm.nih.gov/nuccore/MF872612.1

Field performance of the malaria highly sensitive rapid diagnostic test in a setting of varying malaria transmission

BackgroundThe Gambia has successfully reduced malaria transmission. The human reservoir of infection could further decrease if malaria-infected individuals could be identified by highly sensitive, field-based, diagnostic tools and then treated.MethodsA cross-sectional survey was done at the peak of the 2017 malaria season in 47 Gambian villages. From each village, 100 residents were randomly selected for finger-prick blood samples to detect Plasmodium falciparum infections using highly sensitive rapid diagnostic tests (HS-RDT) and PCR. The sensitivity and specificity of the HS-RDT were estimated (assuming PCR as the gold standard) across varying transmission intensities and in different age groups. A deterministic, age-structured, dynamic model of malaria transmission was used to estimate the impact of mass testing and treatment (MTAT) with HS-RDT in four different scenarios of malaria prevalence by PCR: 5, 15, 30, and 60%, and with seasonal transmission. The impact was compared both to MTAT with conventional RDT and mass drug administration (MDA).ResultsMalaria prevalence by HS-RDT was 15% (570/3798; 95% CI 13.9–16.1). The HS-RDT sensitivity and specificity were 38.4% (191/497, 95% CI 34.2–42.71) and 88.5% (2922/3301; 95% CI 87.4–89.6), respectively. Sensitivity was the highest (50.9%, 95% CI 43.3–58.5%) in high prevalence villages (20–50% by PCR). The model predicted that in very low transmission areas (≤ 5%), three monthly rounds of MTAT with HS-RDT, starting towards the end of the dry season and testing 65 or 85% of the population for 2 consecutive years, would avert 62 or 78% of malaria cases (over 2 years), respectively. The effect of the intervention would be lower in a moderate transmission setting. In all settings, MDA would be superior to MTAT with HS-RDT which would be superior to MTAT with conventional RDT.ConclusionThe HS-RDT’s field sensitivity was modest and varied by transmission intensity. In low to very low transmission areas, three monthly rounds per year of MTAT with HS-RDT at 85% coverage for 2 consecutive years would reduce malaria prevalence to such low levels that additional strategies may achieve elimination. The model prediction would need to be confirmed by cluster-randomized trials.

Is that a real oocyst? Insectary establishment and identification of Plasmodium falciparum oocysts in midguts of Anopheles mosquitoes fed on infected human blood in Tororo, Uganda

BackgroundThe human infectious reservoir for malaria consists of individuals capable of infecting mosquitoes. Oocyst prevalence and density are typical indicators of human infectivity to mosquitoes. However, identification of oocysts is challenging, particularly in areas of low malaria transmission intensity where few individuals may infect mosquitoes, and infected mosquitoes tend to have few oocysts. Here, features that differentiate oocysts from other oocyst-like in mosquito midguts are explained and illustrated. In addition, the establishment and maintenance of infrastructure to perform malaria transmission experiments is described. This work may support other initiatives to set up membrane feeding infrastructure and guide oocyst detection in low transmission settings.MethodsIn 2014, an insectary was developed and equipped in Tororo district, Uganda. A colony of Anopheles gambiae s.s. mosquitoes (Kisumu strain) was initiated to support infectivity experiments from participants enrolled in a large cohort study. Venous blood drawn from participants who were naturally infected with malaria parasites was used for membrane feeding assays, using 60–80 mosquitoes per experiment. Approximately 9–10 days after feeding, mosquitoes were dissected, and midguts were stained in mercurochrome and examined by light microscopy for Plasmodium falciparum oocysts and similar structures. In supportive experiments, different staining procedures were compared using in vitro cultured parasites.ResultsA stable colony of the Kisumu strain of An. gambiae s.s. was achieved, producing 5000–10,000 adult mosquitoes on a weekly basis. Challenges due to temperature fluctuations, mosquito pathogens and pests were successfully overcome. Oocysts were characterized by: presence of malaria pigment, clearly defined edge, round shape within the mosquito midgut or on the peripheral tissue and always attached to the epithelium. The main distinguishing feature between artifacts and mature oocysts was the presence of defined pigment within the oocysts.ConclusionsOocysts may be mistaken for other structures in mosquito midguts. Distinguishing real oocysts from oocyst-like structures may be challenging for inexperienced microscopists due to overlapping features. The characteristics and guidelines outlined here support identification of oocysts and reliable detection at low oocyst densities. Practical advice on sustaining a healthy mosquito colony for feeding experiments is provided. Following the reported optimization, the established infrastructure in Tororo allows assessments of infectivity of naturally infected parasite carriers.

Mass drug administration can be a valuable addition to the malaria elimination toolbox

The Global Technical Strategy 2016–2030 of the World Health Organization (WHO) has the ambitious goal of malaria being eliminated from at least 35 countries by 2030. However, in areas with once-stable malaria transmission, the reservoir of human infection may be intermittently symptomatic or fully silent yet still lead to transmission, posing a serious challenge to elimination. Mass drug administration (MDA), defined as the provision of a therapeutic dose of an effective anti-malarial drug to the entire target population, irrespective of infection status or symptoms, is one strategy to combat the silent human reservoir of infection. MDA is currently recommended by the WHO as a potential strategy for the elimination of Plasmodium falciparum malaria in areas approaching interruption of transmission, given the prerequisites of good access to case management, effective vector control and surveillance, and limited potential for reintroduction. Recent community randomized controlled trials of MDA with dihydroartemisinin–piperaquine, implemented as part of a comprehensive package of interventions, have shown this strategy to be safe and effective in significantly lowering the malaria burden in pre-elimination settings. Here it is argued that effectively implemented MDA should be kept in the elimination toolbox as a potential strategy for P. falciparum elimination in a variety of settings, including islands, appropriate low transmission settings, and in epidemics and complex emergencies. Effectively implemented MDA using an ACT has been shown to be safe, unrelated to the emergence of drug resistance, and may play an important role in sufficiently lowering the malaria burden to allow malaria transmission foci to be more easily identified, and to allow elimination programmes to more feasibly implement case-based surveillance and follow-up activities. To be most impactful and guard against drug resistance, MDA should use an ACT, achieve high programmatic coverage and adherence, be implemented when transmission is lowest in areas of limited risk of immediate parasite reintroduction, and must always be implemented only once good access to case management, high coverage of effective vector control, and strong surveillance have been achieved. If these considerations are taken into account, MDA should prove to be a valuable tool for the malaria elimination toolbox.

From cell culture to cynomolgus macaque: infection models show lineage-specific virulence potential of Coxiella burnetii.

Coxiella burnetii is an obligate intracellular pathogen that causes the zoonotic disease Q fever in humans, which can occur in either an acute or a chronic form with serious complications. The bacterium has a wide host range, including unicellular organisms, invertebrates, birds and mammals, with livestock representing the most significant reservoir for human infections. Cell culture models have been used to decipher the intracellular lifestyle of C. burnetii, and several infection models, including invertebrates, rodents and non-human primates, are being used to investigate host-pathogen interactions and to identify bacterial virulence factors and vaccine candidates. However, none of the models replicate all aspects of human disease. Furthermore, it is becoming evident that C. burnetii isolates belonging to different lineages exhibit differences in their virulence in these models. Here, we compare the advantages and disadvantages of commonly used infection models and summarize currently available data for lineage-specific virulence.

Detection of Escherichia coli, Salmonella spp. and Staphylococcus aureus in Ready-to-Eat Food in Al-Ahsa Province, Saudi Arabia

The study aimed to identify and characterize foodborne- Staphylococcus aureus, Escherichia coli and Salmonella spp. in Al-Ahsa Province, Kingdom of Saudi Arabia (KSA), as potential reservoir of human infection and transmitters of antimicrobial resistance. A total of 90 sandwich samples (consist of minced meat and vegetables) were aseptically collected from fast-food cafeterias. Conventional bacteriological techniques were used to isolate Staph aureus, E. coli and Salmonella spp. For E. coli isolates, molecular analysis was made. Staph aureus was confirmed from 11.11% of specimens of which 30% were MRSA. MRSA were resistant to erythromycin, nitrofurantoin and Trimethoprim/ Sulfamethoxazole. Non-MRSA were resistant to ciprofloxacin and all Staph aureus isolates were sensitive to vancomycin which may be a choice for treatment. At a rate of 5.56%, E. coli was confirmed by conventional techniques and VITEK 2 system; E. coli strain O157: H7 was not identified from the isolates. Molecular analysis indicated that 4 strains belonged to Shiga toxigenic E. coli (STEC) family and one strain was Shigella flexneri. Antimicrobial susceptibility analysis of isolates showed two strains (40%) were extended-spectrum β-lactamases (ESBL) positive that were demonstrated to be susceptible to imipenem but resistant to ciprofloxacin. On the other hand, three strains (60%) were identified as ESBL negative which were susceptible to all tested antibiotics. Salmonella spp. were not isolated from any food specimen in the present study. From ready-to-eat food in Al-Ahsa Province, KSA, Staph aureus MRSA and Non-MRSA were susceptible to vancomycin. Using molecular methods, E. coli and Shig. flexneri were confirmed from food with ESBL positive susceptible to imipenem but resistant to ciprofloxacin.

Assessment of treatment impact on lymphatic filariasis in 13 districts of Benin: progress toward elimination in nine districts despite persistence of transmission in some areas

BackgroundLymphatic filariasis (LF) is still a public health burden in many developing countries. In Benin, a West African country, at least 6.6 million people are at risk for LF. With the goal of eliminating LF by 2020, mass drug administration (MDA) has been scaled-up during the last decade. Currently, 23 districts are believed to have eliminated LF as a public health problem, and 25 other districts are still under treatment. In this study we report the results of the first transmission assessment survey of LF (TAS1) in 13 districts from the second group, which have received at least six rounds of MDA with albendazole and ivermectin.MethodsThe 13 districts were grouped into six evaluation units (EU). In each EU, 30 schools randomly selected by survey sample builder (SSB) software were surveyed. Children aged six and seven were sampled in schools and for each child the Alere™ Filariasis Test Strip test was carried out using finger-prick blood to detect the circulating filarial antigen from Wuchereria bancrofti.ResultsOverall, 9381 children were sampled in 191 schools from the six EU with 47.6% of the children aged six years and 52.4% aged seven years. Five EU passed the assessment, with no positive cases identified. The EU of Ouinhi which grouped the districts of Ouinhi, Cove, Za-Kpota and Zagnanado failed, with 47 positive cases. These cases were clustered in the districts of Ouinhi (n = 20), Za-Kpota (n = 11) and Zagnanado (n = 16). No cases were found in the district of Cove.ConclusionsThe findings of our study indicate that Benin has made important progress towards elimination in most districts evaluated. However, this study also shows that transmission of LF is ongoing in the EU of Ouinhi, part of the Zou department. The MDA strategy needs to be strengthened in order to control the human reservoir of infection in these districts.

Evidence of asymptomatic submicroscopic malaria in low transmission areas in Belaga district, Kapit division, Sarawak, Malaysia

BackgroundMalaysia has declared its aim to eliminate malaria with a goal of achieving zero local transmission by the year 2020. However, targeting the human reservoir of infection, including those with asymptomatic infection is required to achieve malaria elimination. Diagnosing asymptomatic malaria is not as straightforward due to the obvious lack of clinical manifestations and often subpatent level of parasites. Accurate diagnosis of malaria is important for providing realistic estimates of malaria burden and preventing misinformed interventions. Low levels of parasitaemia acts as silent reservoir of transmission thus remains infectious to susceptible mosquito vectors. Hence, the aim of this study is to investigate the prevalence of asymptomatic submicroscopic malaria (SMM) in the District of Belaga, Sarawak.MethodsIn 2013, a total of 1744 dried blood spots (DBS) were obtained from residents of 8 longhouses who appeared healthy. Subsequently, 251 venous blood samples were collected from residents of 2 localities in 2014 based on the highest number of submicroscopic cases from prior findings. Thin and thick blood films were prepared from blood obtained from all participants in this study. Microscopic examination were carried out on all samples and a nested and nested multiplex PCR were performed on samples collected in 2013 and 2014 respectively.ResultsNo malaria parasites were detected in all the Giemsa-stained blood films. However, of the 1744 samples, 29 (1.7%) were positive for Plasmodium vivax by PCR. Additionally, of the 251 samples, the most prevalent mono-infection detected by PCR was Plasmodium falciparum 50 (20%), followed by P. vivax 39 (16%), P. knowlesi 9 (4%), and mixed infections 20 (8%).ConclusionsThis research findings conclude evidence of Plasmodium by PCR, among samples previously undetectable by routine blood film microscopic examination, in local ethnic minority who are clinically healthy. SMM in Belaga district is attributed not only to P. vivax, but also to P. falciparum and P. knowlesi. In complementing efforts of programme managers, there is a need to increase surveillance for SMM nationwide to estimate the degree of SMM that warrant measures to block new transmission of malaria.

Fluoroquinolone-Resistant Salmonella enterica, Campylobacter spp., and Arcobacter butzleri from Local and Imported Poultry Meat in Kumasi, Ghana.

Salmonella and Campylobacter are important gastroenteric pathogens. Arcobacter butzleri is an emerging enteric pathogen. Data on the frequencies of these poultry-associated pathogens on meat products sold in sub-Saharan Africa are scarce. This study aimed to analyze the frequency of Salmonella, Campylobacter, and Arcobacter antibiotic resistance and underlying mechanisms of resistance to fluoroquinolones in locally produced and imported poultry sold in urban Ghana. Chicken meat was collected and cultured on standard media. Bacterial strains were identified by biochemical methods and by mass spectrometry. Antibiotic susceptibility was tested by disk diffusion. Ciprofloxacin-resistant strains were assessed for molecular mechanisms of resistance. Among 200 samples, comprising 34% (n = 68) from the Ghanaian poultry industry and 66% (n = 132) from imports, 9% (n = 17) contained Salmonella, 11% (n = 22) Campylobacter, and 26.5% (n = 53) A. butzleri. Higher overall contamination frequencies were found in local meat. Most common Salmonella serovars identified were Kentucky (n/N = 5/16; 31%) and Poona (n/N = 4/16; 25%). Campylobacter were C. coli (n/N = 10/19; 53%) and C. jejuni (n/N = 9/19; 47%). Resistance to fluoroquinolones was high with 63% (n = 10), 75% (n = 15), and 52% (n = 25) in Salmonella, Campylobacter, and Arcobacter, respectively. A link between Salmonella Kentucky [sequence type (ST) 198] and a ciprofloxacin minimum inhibitory concentration of 16 μg/mL was found. Salmonella Poona-ST308 revealed transferable qnrB2 fluoroquinolone resistance genes. Markedly high frequencies of resistant Salmonella, Campylobacter, and Arcobacter predominant in locally produced meat represent a probable transmission reservoir for human infections. These findings highlight the need for implementation of surveillance systems that focus on food hygiene, use of antibiotics in animal husbandry, and continuous monitoring of the quality of meat products from imports.

First report on the prevalence and subtype distribution of Blastocystis sp. in dairy cattle in Lebanon and assessment of zoonotic transmission

Blastocystis sp. is frequently identified in a wide range of animal hosts, including bovids. Because of its burden and zoonotic potential, this parasite has been sought in domestic cattle from various countries, since this livestock may also represent a possible reservoir of human infection. However, epidemiological data regarding the prevalence and ST distribution of Blastocystis sp. in this animal group is lacking in Lebanon. Therefore, faecal samples were collected from a total of 254 dairy cattle raised on 55 farms located in the North Lebanon region and screened for the presence of the parasite by quantitative real-time PCR. The overall prevalence of Blastocystis sp. was shown to reach 63.4% in cattle livestock. Sequence analysis of positive samples indicated the presence of seven STs, with predominance of ST10 (44.0%) and ST14 (36.8%) and lower proportions of ST2 (8.0%), ST1 (7.2%), ST5 (2.4%), ST3 and ST7 (0.8% each). This survey was the first conducted worldwide reporting ST2 and ST7 in domestic cattle and confirmed that ST10 and ST14 represent cattle-adapted STs in view of their high prevalence. Faecal samples from in-contact dairy farmers and patients hospitalised in the same Lebanese governorate who reported no contact with cattle livestock were also analysed for the presence of Blastocystis sp. The same three STs were identified in both human cohorts, with predominance of ST3, followed either by ST1 or ST2 depending of the group. No other STs, including ST10 or ST14, have been reported. Moreover, even though ST1, ST2 and ST3 were found to be common to dairy cattle and farmers cohorts, only one ST3 isolate showed 100% sequence identity between both hosts. Consequently, the presence and low prevalence of ST1, ST2, ST3, ST5 and ST7 identified herein in domestic cattle, most of which exhibit low host specificity, could be derived from occasional direct exposure to faecal material from human and non-human hosts or by ingestion of contaminated drinking water or food in the enclosure of the farms. Together with the absence of ST10 and ST14 in the human population, these data suggest that cattle play a negligible role as zoonotic reservoirs of Blastocystis sp.

High prevalence of hepatitis E virus infection among domestic pigs in Ibaraki Prefecture, Japan

BackgroundHepatitis E virus (HEV) is prevalent in pigs and may serve as a reservoir for human infection. However, data on HEV infections in pigs in Ibaraki Prefecture, Japan, are limited. Here, we clarified the process and course of HEV in naturally infected pigs. Serum (n = 160) and liver (n = 110) samples were collected from pigs at the slaughterhouse. Furthermore, serum samples were collected from 45 breeding sows and serum and feces samples were collected from 7 piglets once a week (raised until 166 days of age). HEV antigen and antibodies were evaluated, and the genotype was identified based on molecular phylogenetic tree analysis.ResultsThe samples collected from the slaughterhouse revealed that few pigs were HEV carriers but most possessed anti-HEV antibodies. Most breeding sows possessed antibodies, and the piglets excreted HEV on the farm at approximately 10 weeks of age. One pig was initially infected, and in a few weeks, the other pigs living in the same sty became infected.ConclusionsMost pigs in Ibaraki Prefecture were with HEV. On the farm, most piglets were infected with HEV by the time they reached slaughter age. We confirmed that HEV infection is successively transmitted among piglets living in the same sty.

Efficacy of an Adjuvanted Middle East Respiratory Syndrome Coronavirus Spike Protein Vaccine in Dromedary Camels and Alpacas.

MERS-CoV is present in dromedary camels throughout the Middle East and Africa. Dromedary camels are the primary zoonotic reservoir for human infections. Interruption of the zoonotic transmission chain from camels to humans, therefore, may be an effective strategy to control the ongoing MERS-CoV outbreak. Here we show that vaccination with an adjuvanted MERS-CoV Spike protein subunit vaccine confers complete protection from MERS-CoV disease in alpaca and results in reduced and delayed viral shedding in the upper airways of dromedary camels. Protection in alpaca correlates with high serum neutralizing antibody titers. Lower titers of serum neutralizing antibodies correlate with delayed and significantly reduced shedding in the nasal turbinates of dromedary camels. Together, these data indicate that induction of robust neutralizing humoral immune responses by vaccination of naïve animals reduces shedding that potentially could diminish the risk of zoonotic transmission.

Mechanisms underlying Enterohaemorrhagic Escherichia coli O157: H7 manipulation of the bovine cellular immune response

Enterohaemorrhagic Escherichia coli (EHEC) O157:H7 can cause haemorrhagic diarrhoea and potentially fatal renal failure in humans. Ruminants are considered the primary reservoir for human infection. Studies investigating the response of cattle to colonization generally focus on humoral immunity, leaving the role of cellular immunity unclear. These bacteria colonise their host by tight attachment to the epithelium, using a type three secretion system to inject a cocktail of effectors into the host cell. Injected effectors manipulate the innate response in several ways to promote bacterial persistence. Transcriptional profiling of responses at the terminal rectum, the primary site of colonisation in cattle, reveals a bias towards a T-helper type 1 response, indicating that cellular immunity may be involved in bacterial clearance. Mathematical modelling also indicates that injected effectors have a reduced human MHC-I epitope density, whilst structural bacterial proteins do not. This implies that host cellular immune responses target injected effectors and have exerted selective pressure on their evolution. My on-going research is examining the expression of MHC-I on the surface of cultured bovine epithelial cells following infection with E. coli O157. Initial results demonstrate a decrease in MHC-I surface expression during EHEC O157 colonisation after three hours. The basis of this reduction is being investigated using defined mutants in type three secretion genes. The longer-term objective is to use peptide elution and mass spectrometry to examine the presentation of effector protein peptides and determine whether E. coli O157 has evolved to interfere with this process.

English

A cross-sectional study was conducted from December 2016 to May 2017 in and around Sebeta town with the aim of assessing the prevalence of mastitis, isolation of aerobic bacterial and fungal causal agents and assessing antimicrobial resistance pattern of isolated Staphylococcus species in dairy cows. From a total of 383 dairy cows, 220 (57.4%) were found to be positive for mastitis of which 10.4% were affected by clinical mastitis and 47% by subclinical mastitis. Mastitis was more likely to occur in cows above 8 years of age (OR = 16.9, 95% CI = 7.8 - 36.00) and in those cows washed by hand before milking (OR =7.8, 95% CI = 4.2-14.6) as compared to those subjected to udder washing and drying using towels. Staphylococcus aureus was the most frequently isolated bacterial species (25%) followed by Streptococcus agalactiae (12.3%) and coagulase negative Staphylococcus species (10.5%). Yarrowia lipolytica (10.9%) and Candida etchellsii (7.3%) were the major yeast species isolated, while Aspergillus (6.8%), Mucor (5.9%), Penicillium (3.6%) and Fusarium (3.6%) were the major filamentous fungi species identified from the cultured milk samples. The results of antimicrobial susceptibility testing revealed that the isolated Staphylococcus species were highly resistant to penicillin G (93.1%) and oxytetracycline (79.3%) but were susceptible to vancomycin (100%), sulphamethoxazole/trimethoprim (96.6%), ampicillin (89.7%) and erythromycin (86.2%). It could be concluded that bovine mastitis is a major challenge to the dairy producers in and around Sebeta towns. Appropriate control and preventative measures must be instituted and dairy farmers and workers must be trained on proper milking and hygiene practices in order to reduce the prevalence of mastitis in this region. The penicillin resistant S. aureus could be a source of serious infection in humans as well and hence comprehensive studies including molecular characteristics of drug resistance gene of S. aureus especially of methicillin-resistant should be conducted as farm animals, primarily dairy cattle might serves as a reservoir of infection for humans. In 2% of the cases, only fungal species were identified as causes of mastitis, hence further investigation regarding their pathogenicity and contribution to bovine mastitis is needed. Key words: Antimicrobial resistance, bovine mastitis, bacteria, Ethiopia, fungus, prevalence, Sebeta.

A multiplex assay for the sensitive detection and quantification of male and female Plasmodium falciparum gametocytes

BackgroundThe transmission of malaria to mosquitoes depends on the presence of gametocytes that circulate in the peripheral blood of infected human hosts. Sensitive estimates of the densities of female gametocytes (FG) and male gametocytes (MG) may allow the prediction of infectivity to mosquitoes and thus a molecular estimate of the human infectious reservoir for transmission.MethodsA novel multiplex qRT-PCR assay with intron-spanning primers was developed for the parallel quantification of FG and MG. CCp4 (PF3D7_0903800) transcripts specific for FG and PfMGET (PF3D7_1469900) transcripts specific for MG were quantified in total nucleic acids. The assay was validated on sex-sorted gametocytes from culture material and on samples from clinical trials with gametocytocidal drugs. Synthetic RNA standards were generated for the two targets genes and calibrated against known gametocyte quantities.ResultsThe limit of detection was determined at 0.1 male and 0.1 female gametocyte/µL, which was equal to the limit of quantification (LOQ) for MG, while the LOQ for FG was 1 FG/µL. Results from previously reported clinical trials that used separate gametocyte qRT-PCR assays for FG (targeting Pfs25) and MG (targeting PfMGET) were reproduced with the multiplex assay. High levels of agreement between separate assays and the multiplex approach were observed (R2 = 0.9473, 95% CI 0.9314–0.9632, for FG measured by transcript levels of Pfs25 in qRT-PCR or CCp4 in multiplex; R2 = 0.8869, 95% CI 0.8541–0.9197, for MG measured by PfMGET in either single or multiplex qRT-PCR). FG and MG transcripts were detected in pure ring stage parasites at 10,000- and 100,000-fold reduced frequency for CCp4 and PfMGET, respectively. The CCp4 and PfMGET transcripts were equally stable under suboptimal storage conditions.ConclusionsGametocyte densities and their sex ratios can be determined in the presented one-step multiplex assay with higher throughput than single assays. The interpretation of low gametocyte densities at asexual parasite densities above 1000 parasites/µL requires caution to avoid false positive gametocyte signals from spurious transcript levels in ring stage parasites.