I have earned a bachelor degree (DVM) in Veterinary Medicine in 2005, since then I have established my research career in the field of Parasite Disease Epidemiology and Antiparasitic Drug Resistance. Therefore I pursue a MSc degree in Veterinary Parasitology, MSc in Molecular Biology, and MSc in Biomedicine from Sweden. I did my PhD in the field of Molecular Parasitology from the School of Veterinary Medicine, University of Calgary, Canada. Since defending my thesis in May 2015, I worked at the School of Veterinary Studies, Edinburgh, Scotland and recently moved to the School of Veterinary Medicine, Surrey, England.
Areas of specialism
University roles and responsibilities
- VMS2009- Concept of Infectious Diseases (BVM&S)
- VMS3009- Veterinary Research Project (BVM&S)
- VMS1007- Structure and Function-5: Reproductive System (BVM&S)
- VMS2010- Animals in Society-3 Module (BVM&S)
- VMS3011-Veterinary Medicine-2 Module (BVM&S)
I took a lead for the development of novel high throughput ‘haemoprotobiome’ tool and applied it to quantify the protozoan infection in livestock and humans. I develop the novel ‘tremabiome’ tool for the identification of fluke parasites and the distribution of trematode infection in their definitive hosts. I have modified the ‘nemabiome’ tool, and develop the pyrosequencing tools to quantify the proportions of nematode parasites in various co-infection studies. Sensitive diagnostic tools are needed for better-informed disease surveillance and evaluating parasite responses to drug treatments.
Another key component of my work is to develop a novel multiplicity of infection (MOI) based genetic models to explain how the infection raises and transmitted in protozoan and fluke parasites of livestock. The information will contribute to the understanding of host-parasite relationships, influences of vectors and, or intermediate hosts, and climate change on the epidemiology of parasitic diseases, thereby aiding in the development of parasite control strategies.
I develop expertise to study the antigenic diversity in livestock and human parasites. The overall aim is to define the antigenic relationship between parasite populations for the foundation of future validation studies of antigenicity and immunogenicity of vaccine candidate loci capable of providing cross-protection against different parasite strains at the regional level.
I have led a research project to study the antiparasitic drug resistance in the nematode and protozoan parasites of livestock and humans. I have demonstrated the selective sweep based genetic model to explain how the antiparasitic drug resistance mutations could develop, allowing different predictions to be made about the consequences of selection pressures, fitness costs, and gene flow in the emergence and spread of resistance alleles in parasite populations. I worked on the construction of a genetic linkage map and XQTL analysis to aid in the identification of antiparasitic drug resistance loci that help to improve long-term sustainable resistance control measures.
Postgraduate research supervision
I have supervised several PhD, MSc/VetMed Resident and undergraduate (BVM&S, BSC, MBChb) students in UK, in addition to overseas PhD and MSc students in Pakistan, Sweden, and France. To date, 4 of 6 PhDs, 9 of 10 MSc/VetMed Resident and 8 of 10 undergraduate students have successfully completed their degrees. I work with my students to develop their research projects, while concurrently trained them in research methods. My students have taken the research projects on the number of different topics including parasite disease epidemiology, population genetics of antiparasitic drug resistance, novel molecular diagnostic tools, bioinformatics, and phylogenetic modelling. They first begin to read broadly across relevant literature and identify a research project of interest and considers the best possible dataset methods needed to tackle the research questions. Once they become comfortable with research practices, our focus shifts to develop analytical methods and to the clear presentation of their research findings, with a goal of presenting their work at a conference or co-authorship. To date, 4 of 6 PhDs, and 8 of 10 MSc/VetMed Resident students are successful for the co-authorship in high-quality peer-reviewed scientific journals. In this professional environment, students are immersed in a research community where research experiences are intensified, but the processes and approaches of scientific understanding, careful analysis, and clear communication should be maintained.
I contribute to the teaching of the parasitology module (BVM&S). A crucial component of this course is to demonstrate both the life cycles of parasites and the significance of parasites in their ecosystems. For example, my students have successfully explored the features of arthropods parasites in combination with clinical information. They recognise the structure and feeding mechanisms of tick, lice, flea, and flies. They can identify the ciliate and flagellate protozoa by their type of movement and distinguish the protozoan parasites in the stained smear. They learn the nematode egg morphology with faecal egg count methods. They examine the specimens of liver and rumens having fluke infestation and the specimens of lungs having tapeworms cyst.
I teach veterinary research project model for BVM&S students at the School of Veterinary Medicine, University of Surrey. In this module, my students develop their research skills, while concurrently trained in research methods.
I teach in the structure of the reproductive system module at the School of Veterinary Medicine, University of Surrey BVM&S students. In this module, I contribute to the lambing, stomach tubbing, castration and tail docking simulation using cadaver neonatal lambs. I also teach the animal in society module at the School of Veterinary Medicine, University of Surrey BVM&S students. In this module, I contribute to the safe animal handling and restraining, TPRs, intravenous, intramuscular and subcutaneous injection learning.
I also contribute to the laboratory tools for the molecular biology module (MSc). This course aims to introduce the basic techniques required to work with DNA. In this course, my students start with pathogen-infected cells and extract the DNA followed by PCR amplification. They clone the PCR product in the plasmid and analysed by restriction digestion to ensure the cloning has been successful, before submitting the DNA for sequencing. They amplify the DNA by real-time quantitative PCR (qPCR) and assess the pathogen load in samples.
The benzimidazoles are one of the most important broad-spectrum anthelmintic drug classes for parasitic nematode control in domestic animals and humans. They have been widely used in livestock, particularly in small ruminants for over 40 years. This has resulted in widespread resistance in small ruminant gastrointestinal nematode parasite species, especially Haemonchus contortus. Benzimidazole resistance mutations have also been reported in Haemonchus placei, but only at low frequencies, suggesting resistance is at a much earlier stage of emergence than is the case for H. contortus. Here, we investigate the haplotype diversity of isotype-1 β-tubulin benzimidazole resistance mutations and the population genetic structure of H. contortus and H. placei populations from sheep and cattle from the southern USA. Microsatellite genotyping revealed a low level of genetic differentiation in six H.placei and seven H. contortus populations examined. This is consistent with several previous studies from other regions, mainly in H. contortus, supporting a model of high gene flow between parasite populations. There was a single F200Y(TAC) haplotype present in all six H. placei populations across Georgia, Florida and Arkansas. In contrast, there were at least two different F200Y(TAC) haplotypes (up to four) and two different F167Y(TAC) haplotypes across the seven H. contortus populations studied. These results provide further evidence to support a model for benzimidazole resistance in Haemonchus spp, in which resistance mutations arise from a single, or the small number of locations, in a region during the early phases of emergence, and subsequently spread due to animal movement.
The large stomach worm, Haemonchus, commonly known as the barber’s pole worm, is a blood sucking nematode found in the abomasa of small and large ruminants. Allele-specific amplification of the rDNA internal transcribed spacer-2 (ITS-2) sequences was performed from the total of 78 individual adult worms to screen Haemonchus placei at species level, which is the significant diagnostic tool to identify this major economically important species. Further full sequences analysis of the ITS-2 region revealed that there are 4 sites shows intraspecific variations at position 65, 111, 125 and 148. For instance this study is the first documented report of intraspecific genetic variations in the rDNA ITS-2 sequences of H. placei from cattle in Pakistan and the results shows that H. placei is genetically different from the isolates studied previously. However, detailed and large size samples strategy will be required to identify the co-infection and interspecies hybridization between H. placei and Haemonchus contortus in cattle.
Background The impact of drug selection pressure on the overall genetic diversity of parasitic nematode populations in the field is poorly understood. In this study, we address this issue for the small ruminant parasite Haemonchus contortus in the Punjab, Pakistan. This region provides an opportunity to compare H. contortus populations that have been subjected to a prolonged period of frequent benzimidazole drug treatments on government farms with parasite populations that have been exposed to little or no drug treatment in neighbouring pastoral herds. Methods Adult H. contortus worms were collected from the abomasa of small ruminants from three government farms frequently using benzimidazole drugs, and closed to animal movement, for over 30 years and also from from eighteen pastoral herds subject to minimal drug selection. The frequency of three known benzimidazole resistance associated mutations was determined in each parasite population. For the seven parasite populations in which resistance mutations were found, the diversity, geographical distribution and phylogenetic relationships of isotype-1 β-tubulin benzimidazole resistance haplotypes were determined. In addition, the genetic diversity of the parasite populations on the three government farms were compared with those from four pastoral herds. Results The F200Y (TAC) resistance mutation was present at a very high frequency in H. contortus populations from government herds, but not from pastoral herds, consistent with their respective drug selection histories. Population genetic analysis, using a panel of microsatellite markers, revealed that there was little genetic differentiation among the parasite populations with no significant difference in the overall genetic diversity between government and pastoral herds. In addition, sequence analysis of the isotype-1 β-tubulin locus revealed multiple F200Y (TAC) haplotypes demonstrating soft selective sweeps even in government herds with little or no contemporary parasite migration. Conclusions The results suggest that, although the frequent drug treatment used on government farms has selected for a high frequency of benzimidazole resistance mutations, there has been little or no reduction in the overall genetic diversity of the selected parasite populations.
Insular wildlife populations provide opportunities to examine biological questions in systems that are relatively closed and potentially tractable, striking examples being the long-term studies of ecology and evolution in the red deer and feral sheep populations on the Hebridean islands of Rum and St Kilda. In the case of parasitology, Understanding of parasitic infections insular wildlife populations in conjunction with knowledge of their origins has the potential to add a fresh perspective to disease control in humans and domestic animals. In the case of parasitology, understanding infections of insular wildlife populations, in conjunction with knowledge of their origins, has the potential to add a fresh perspective to disease control in humans and domestic animals. With this in mind, gross and molecular examination for the presence of cyclophyllidean tapeworms was performed on the viscera and rectal contents of 17 preserved specimens of Apodemus sylvaticus field mice and on the naturally voided faeces of a further four mice on the remote archipelago of St Kilda. Molecular speciation of hexacanth embryos extracted from the faeces of two mice, using nucleotide sequence analysis of the ribosomal cytochrome c-oxidase subunit-1, confirmed infection with Hymenolepis hibernia. Phylogenetic analysis showed that these were genetically distinct from Hymenolepis diminuta, previously reported in the insular A. sylvaticus mice, and from other published H. hibernia haplotypes. There was insufficient hymenolepidid tapeworm phylogeographic variation to resolve the origins of the co-evolved St Kilda mice, primarily due to a lack of published H. hibernia Cox-1 sequence data across the parasite's geographical range. Nevertheless, the Maximum Likelihood haplotype tree shows the potential for molecular parasitology to resolve a host-parasite relationship once more data become available. Morphological diagnostic features of zoonotic H. hibernia eggs are also described. [Display omitted] •First identification of Hymenolepis hibernia in Apodemus sylvaticus hirtensis mice.•Molecular speciation of hexocanth eggs.•Morphological description of Hymenolepis hibernia eggs.•Maximum likelihood hymenolepidid tapeworm haplotype tree.•Consideration of parasite origins and those of co-evolved hosts.
Canine parvoviruses (CPV) exist as antigenic variants with varying frequencies and genetic variabilities across the globe. Given the endemicity and high prevalence in Pakistan, we characterized the CPVs originating from dogs-population to elucidate viral diversity and evolution. Fecal samples from clinically diseased pups (n = 17) of different breeds and age (2-6 months) were processed for hemagglutination assay (HA), and later for partial amplification of VP2 gene sequence and amino acid analysis. A total of 11 samples (64.71%) were found positive both in hemagglutination and PCR assays. Phylogenetic and evolutionary analysis demonstrated higher genetic heterogeneity in studied strains and constituted seven clusters within the CPV-2a group, however, they shared high level of identity with Chinese strains. Further studies are necessary to elucidate genetic analysis and epidemiology of CPV variants across a wide geographical area of the country.
Haemonchus populations were collected from cattle from mid-western and eastern Southern US (four and six populations, respectively) to determine the relative prevalence of Haemonchus contortus and Haemonchus placei and the frequency of the three isotype-1 β-tubulin polymorphisms associated with benzimidazole resistance. A minimum of 32 individual adult worms were genotyped at position 24 of the rDNA ITS-2 for each population to determine species identity (296 worms in total). One population from Georgia was identified as 100% H. contortus with the remaining nine populations identified as 100% H. placei. For the H. contortus population, 29 out of 32 worms carried the P200Y (TAC) isotype-1 β-tubulin and 2 out of 32 worms carried the P167Y (TAC) benzimidazole resistance associated polymorphisms respectively. For H. placei, six out of the nine populations contained the P200Y (TAC) isotype-1 β-tubulin benzimidazole resistance associated polymorphism at low frequency (between 1.6% and 9.4%) with no resistance associated polymorphisms being identified at the P198 and P167 codons. This is the first report of the P200Y (TAC) isotype-1 β-tubulin benzimidazole resistance associated polymorphism in H. placei. The presence of this mutation in multiple independent H. placei populations indicates the risk of resistance emerging in this parasite should benzimidazoles be intensively used for parasite control in US cattle.
Background Ivermectin is an endectocide against many parasites. Though being a macrocyclic lactone, its activity against bacteria has been less known, possibly due to the fact that micromolar concentrations at tissue levels are required to achieve a therapeutic effect. Among pathogenic bacteria of major medical significance, Staphylococcus aureus cause a number of diseases in a wide variety of hosts including humans and animals. It has been attributed as one of the most pathogenic organisms. The emergence of methicillin resistance has made the treatment of S. aureus even more difficult as it is now resistant to most of the available antibiotics. Thus, search for alternate anti-staphylococcal agents requires immediate attention. Methods Twenty-one clinical isolates of S. aureus were isolated from bovine milk collected from Lahore and Faisalabad Pakistan. Different anthelmintics including levamisole, albendazole and ivermectin were tested against S. aureus to determine their minimum inhibitory concentrations. This was followed-up by growth curve analysis, spot assay and time-kill kinetics. Results The results showed that ivermectin but not levamisole or albendazole exhibited a potent anti-staphylococcal activity at the concentrations of 6.25 and 12.5 μg/ml against two isolates. Interestingly, one of the isolate was sensitive while the other was resistant to methicillin/cefoxitin. Conclusions Our novel findings indicate that ivermectin has an anti-bacterial effect against certain S. aureus isolates. However, to comprehend why ivermectin did not inhibit the growth of all Staphylococci needs further investigation. Nevertheless, we have extended the broad range of known pharmacological effects of ivermectin. As pharmacology and toxicology of ivermectin are well known, its further development as an anti-staphylococcal agent is potentially appealing.
Toxoplasma gondii is an intracellular parasite, which infects human and animals by ingestion of tissue cyst, raw or undercooked meat or oocyst from soil, vegetables, fruits, water, soil and food contaminated by cat faeces or by transmission through the placenta, milk and blood transfusion. Seropositivity levels vary widely among different regions of the globe and according to sociocultural habits, geographic factors; climate and transmission routes and typically rise with age. In view of the worldwide importance of T. gondii, a study was conducted to determine the prevalence of T. gondii antibody in camels by using Toxoplasma Latex Test Kit. The overall prevalence of T. gondii infection in camels was recorded as 10%. Two camels were found seropositive at 1:16 dilution showing residual or nonspecific immunity, five camels were found seropositive at 1:128 showing acquired or evolving immunity, whereas three camels were positive at antibody titre of 1:256 giving an evidence of present infection. It was also noted that seropositivity of T. gondii in camels was higher in age group from 6–10 years; infection was higher in female camels having abortion history.
Genetic hybridisation between parasitic nematode species has potentially important consequences. It could lead to the introgression of genes between species including those involved in pathogenicity, host specificity, transmission and drug resistance. It could also complicate diagnosis and control. However, there are few compelling examples of its occurrence in parasites in the field. Haemonchus contortus and Haemonchus placei are two closely related parasitic nematode species that predominantly infect small ruminants and cattle, respectively. They are capable of experimental hybridisation when adult worms of each species are transplanted into the same individual host. Given that co-infection occurs in both small ruminants and cattle, there is potential for hybridisation in the field. However, this has not been definitively demonstrated and its extent is unknown. We investigated the occurrence of co-infection and interspecies hybridisation in H. contortus and H. placei in field populations from small ruminants from Pakistan and southern India using a number of independent genetic markers. Haemonchus contortus and H. placei co-infections were common in Pakistan but not in southern India where H. placei appeared to be absent in small ruminant hosts. In the former region, a number of worms were identified that were heterozygous for fixed, species-specific rDNA internal transcribed spacer 2 (ITS-2) single nucleotide polymorphisms. Genotyping of these ITS-2 heterozygotes with an additional four nuclear markers conclusively demonstrated them to be F1 interspecies hybrids. Mitochondrial NADH dehydrogenase subunit 4 haplotype analysis demonstrated that four of the hybrid worms had a H. placei maternal parent and one had a H. contortus maternal parent showing that hybridisation could occur in either direction. Interestingly, one of these hybrids contained an H. contortus isotype-1 β-tubulin benzimidazole resistance allele, suggesting there is a potential for interspecies introgression of drug resistance loci. We believe this is the first definitive genetic evidence of hybridisation between H. contortus and H. placei in the field and represents the most comprehensive genetic evidence of F1 hybrids between any human or livestock parasitic nematode species to date. Further, it suggests that interspecies transmission of anthelmintic resistance mutations warrants further investigation.
Since the first report of infectious bursal disease in Pakistan in 1987, outbreaks have been common even in vaccinated flocks. Despite appropriate administration of vaccines, concerns arise if the circulating strains are different from the ones used in the vaccine. Here, we sequenced the hypervariable region (HVR) of the VP2 gene of circulating strains of infectious bursal disease virus (IBDV) originating from outbreaks (n = 4) in broiler flocks in Pakistan. Nucleotide sequencing followed by phylogeny and deduced amino acid sequence analysis showed the circulating strains to be very virulent (vv) and identified characteristic residues at position 222 (A), 242 (I), 256 (I), 294 (I) and 299 (S). In addition, a substitution at positions 221 (Q→H) was found to be exclusive to Pakistani strains in our analysis, although a larger dataset is required to confirm this finding. Compared to vaccine strains that are commonly used in Pakistan, substitution mutations were found at key amino acid positions in VP2 that may be responsible for potential changes in neutralization epitopes and vaccine failure.
It is important to understand how anthelmintic drug resistance mutations arise and spread in order to determine appropriate mitigation strategies. We hypothesised that a molecular genetic study of Haemonchus contortus in southern India, a region where resistance may be less advanced than in western Europe and North America, might provide some important insights into the origin and spread of anthelmintic resistance. The F200Y (TAC) isotype-1 β-tubulin benzimidazole resistance mutation is common in H. contortus throughout the world and the F167Y (TAC) and E198A (GCA) mutations, although less common, have been reported in a number of different countries. We have investigated the haplotypic diversity and phylogenetic relationship of isotype-1 β-tubulin benzimidazole resistance alleles for 23 H. contortus populations from small ruminants across southern India. The F200Y (TAC) mutation was most common, being detected in 18/23 populations at frequencies between 9% and 84% and the E198A (GCA) mutation was also detected in 8/23 populations at frequencies between 8% and 18%. The F167Y (TAC) mutation was not detected in any of the 23 populations. Phylogenetic haplotype network analysis suggested that the F200Y (TAC) mutation has arisen multiple independent times in the region with at least three independent origins of resistance alleles across the populations surveyed. In contrast, the E198A (GCA) mutation was present on a single haplotype which, given the high level of haplotypic diversity of the susceptible alleles in the region, suggests this particular mutation has spread from a single origin, likely by anthropogenic animal movement. Population genetic analysis of 12 of the H. contortus populations, using a panel of eight microsatellite markers, revealed extremely low genetic differentiation between populations, consistent with the hypothesis of high gene flow among sites. Additionally, there was no significant genetic differentiation between H. contortus taken from sheep and goats which is consistent with H. contortus populations being freely shared between these two different hosts. Overall, we believe these results provide the first clear genetic evidence for the spread of an anthelmintic resistance mutation to multiple different locations from a single origin.
The parasitic nematode Haemonchus contortus is an economically and clinically important pathogen of small ruminants, and a model system for understanding the mechanisms and evolution of traits such as anthelmintic resistance. Anthelmintic resistance is widespread and is a major threat to the sustainability of livestock agriculture globally; however, little is known about the genome architecture and parameters such as recombination that will ultimately influence the rate at which resistance may evolve and spread. Here, we performed a genetic cross between two divergent strains of H. contortus, and subsequently used whole-genome resequencing of a female worm and her brood to identify the distribution of genome-wide variation that characterizes these strains. Using a novel bioinformatic approach to identify variants that segregate as expected in a pseudotestcross, we characterized linkage groups and estimated genetic distances between markers to generate a chromosome-scale F1 genetic map. We exploited this map to reveal the recombination landscape, the first for any helminth species, demonstrating extensive variation in recombination rate within and between chromosomes. Analyses of these data also revealed the extent of polyandry, whereby at least eight males were found to have contributed to the genetic variation of the progeny analyzed. Triploid offspring were also identified, which we hypothesize are the result of nondisjunction during female meiosis or polyspermy. These results expand our knowledge of the genetics of parasitic helminths and the unusual life-history of H. contortus, and enhance ongoing efforts to understand the genetic basis of resistance to the drugs used to control these worms and for related species that infect livestock and humans throughout the world. This study also demonstrates the feasibility of using whole-genome resequencing data to directly construct a genetic map in a single generation cross from a noninbred nonmodel organism with a complex lifecycle.
More than 70 species of the family Paramphistomatidae, have been identified in ruminants in different parts of the world. Most are pathogenic, causing amphistomosis. Adult flukes of this Family have a predilection for the rumen, liver or bile duct of ruminants where they may cause damage to the epithelium. Identification of adult paramphistomes to the species level based on morphology alone requires specialized knowledge, whereas, molecular genetic marker analysis is more precise and transferable. In the present study, we performed both morphological and molecular characterization of fifteen adult flukes collected from the liver of domesticated buffalo in the Punjab province of Pakistan. The morphology of five of these flukes was examined in detail and on this basis these were identified as either Explanatum explanatum or Explanatum bathycotyle. PCR and sequencing of the ITS-2 rDNA region from these 5 flukes, plus 10 others, revealed a single haplotype in all cases. This differed by just a single nucleotide polymorphism from a previously described E. explanatum ITS-2 rDNA sequence. Phylogenetic comparison of these E. explanatum ITS2-rDNA sequences with those from other Paramphistomatidae, Fasciola and Dicrocoelium species was performed to assess within and between species variation and validate the use of ITS-2 rDNA as a robust species-specific marker for E. explanatum. This work provides a validated species-specific marker of E. explanatum and the first report of this parasite species from Pakistan.
Fascioliasis is an important disease affecting livestock, with great costs to producers worldwide. It has also become a serious issue for human populations in some endemic areas as an emerging zoonotic infection. There are two Fasciola species of liver fluke responsible for this disease, which occur worldwide, Fasciola hepatica and Fasciola gigantica. Identifying these two species on the basis of adult or egg morphology requires specialist knowledge due to the similarity of characters, and may misidentify putative intermediate or hybrid forms. In this study we sequenced the internal transcribed spacer 2 (ITS-2) rDNA of liver flukes collected from multiple species of hosts from seven localities in the Punjab and Baluchistan provinces of Pakistan, to determine the distribution of these two species. All 46 flukes processed in this study, collected from seven sites, showed the rDNA ITS-2 genotype corresponding to F. gigantica, contradicting previous reports, based on adult and egg morphology, that both species are present in Pakistan, with F. hepatica being the more common.
There is a need for improved methods for the study of the impacts of climatic and livestock management change on the epidemiology of production-limiting helminth parasitic diseases. In this study we report the application of molecular methods to describe the natural history of the small lancet fluke, Dicrocoelium dendriticum on Machair pastures on the Inner Hebridean Isle of Coll. Our results build upon those of the only previous historic field study of D. dendriticum in the British Isles that had been undertaken on the same study site. We demonstrate the value of combining conventional parasitological methods with PCR amplification of a mitochondrial DNA fragment for the detection of D. dendriticum in ants and snails, and PCR amplification of ITS2 and 28S ribosomal DNA fragments to support the species identity of the intermediate hosts, to improving understanding of the epidemiology of D. dendriticum. We report the presence of D. dendriticum infection in cattle, sheep and rabbits grazing on Machair pastures. D. dendriticum infection was identified in a high percentage of the snails, identified as Cochlicella acuta and Cernuella virgata, and in a high percentage of Formica fusca and Myrmica ruginoides ants that were collected from, or clinging to, the tops of flowers. We have identified the involvement of different intermediate host species and higher prevalences of snail and ant infection than previously reported, in part reflecting differences between the sensitivity and specificity of morphological and molecular speciation methods. Overall, our results highlight the complex life history of dicrocoeliosis and illustrate the parasite’s generalist host strategy that confers potential to exploit new niches created by climatic change or grazing management for habitat conservation.
•58R and 173L single mutants and 58R/117N double mutants are present on a single lineage; suggesting a single origin of these mutations.•117N mutant is present on two separate lineages suggesting that there are multiple origins of this mutation.•Single mutation of 117N and double mutations of 58R/117N arise commonly, whereas the single mutation of 173L are less common. Pyrimethamine was first introduced for the treatment of malaria in Asia and Africa during the early 1980s, replacing chloroquine, and has become the first line of drugs in many countries. In recent years, development of pyrimethamine resistance in Plasmodium vivax has become a barrier to effective malaria control strategies. Here, we describe the use of meta-barcoded deep amplicon sequencing technology to assess the evolutionary origin of pyrimethamine resistance by analysing the flanking region of dihydrofolate reductase (dhfr) locus. The genetic modelling suggests that 58R and 173L single mutants and 58R/117N double mutants are present on a single lineage; suggesting a single origin of these mutations. The triple mutants (57L/58R/117N, 58R/61M/117N and 58R/117N/173L) share the lineage of 58R/117N, suggesting a common origin. In contrast, the 117N mutant is present on two separate lineages suggesting that there are multiple origins of this mutation. We characterised the allele frequency of the P. vivax dhfr locus. Our results support the view that the single mutation of 117N and double mutations of 58R/117N arise commonly, whereas the single mutation of 173L and triple mutations of 57L/58R/117N, 58R/61M/117N and 58R/117N/173L are less common. Our work will help to inform mitigation strategies for pyrimethamine resistance in P. vivax.
Our current understanding of differences in the epidemiology of gastrointestinal nematode (GIN) species in co-grazed sheep and goats is inadequate with reference to the development of sustainable control strategies. The next-generation metabarcoding sequencing method referred to as the ‘nemabiome’ allows some of these differences to be explored to describe the intensity of co-infecting GIN species. We applied this platform to study sheep and goats that were co-grazed on Guinea grass pasture in northeastern Brazil. Co-grazed goats and sheep were treated with a monepantel anthelmintic, then exposed to the same gastrointestinal nematode species. Overall, there were differences in the prevalence of GIN species identified in the sheep and goats; Trichostrongylus colubriformis and Teladorsagia circumcincta predominated in goat kids, while Haemonchus contortus predominated in adult does, ewes and lambs once burdens became re-established after anthelmintic treatment. Description of the pattern of re-infection following anthelmintic treatment was prevented by the unpredicted poor efficacy of 2.5 mg/kg and 5 mg/kg, respectively, of monepantel against O. columbianum and T. circumcincta in lambs, and T. circumcincta adult does. Differences in drug efficacy between host age and species groups may be important when considering sustainable GIN control strategies for co-grazed animals. The aggregated FECs of the adult does and goat kids representing re-established GIN burdens, were higher than those of the co-grazed adult ewes and lambs. This implies that there are inherent differences in GIN species adaptation to the two naïve small ruminant host species, and shows the need for better understanding of the factors giving rise to this situation associated with exposure to infective larvae and host responses. At the start of the study, the adult does were co-infected with several GIN species, with the highest intensity of T. circumcincta, contrasting with the situation in the adult ewes, in which H. contortus predominated. However, once burdens became re-established after treatment, H. contortus predominated in both adult does and ewes. This demonstrates the potential for host burdens of H. contortus to establish and predominate after anthelmintic treatment when burdens of co-infecting GIN species are low.
•Summary of the methods currently available for the detection of antimalarial drug resistance within blood samples of human patients.•No single method is perfect for every application, many newly developed methods give promise for a more reliable and efficient characterisation of a Plasmodium resistance.•Deeper understanding of the evolutionary and spaciotemporal dynamics of this disease. Malaria is the world's deadliest parasitic disease. Great progress has been made in the fight against malaria over the past two decades, but this has recently begun to plateau, in part due to the global development of antimalarial drug resistance. The ability to track drug resistance is necessary to achieve progress in treatment, disease surveillance and epidemiology, which has prompted the development of advanced diagnostic methods. These new methods provide unprecedented access to information that can help to guide public health policies. Development of new technologies increases the potential for high throughput and reduced costs of diagnostic tests; improving the accessibility of tools to investigate the forces driving disease dynamics and, ultimately, clinical outcomes for malaria patients and public health. This literature review provides a summary of the methods currently available for the detection of antimalarial drug resistance from the examination of patients’ blood samples. While no single method is perfect for every application, many of the newly developed methods give promise for more reliable and efficient characterisation of Plasmodium resistance in a range of settings. By exploiting the strengths of the tools available, we can develop a deeper understanding of the evolutionary and spatiotemporal dynamics of this disease. This will translate into more effective disease control, better-informed policy, and more timely and successful treatment for malaria patients.
Among helminthes, the trichostrongyloidea consists of most significant parasites of ruminants, which are posing significant threats to livestock productivity, health and well-being. This brief commentary provides an overview of this important group of parasite to showcase the crucial roles these play in hosts.
The prevalence of C. daubneyi infection in the United Kingdom has increased, but despite the potential for rumen flukes to cause production loss in ruminant livestock, understanding of their emergence and spread is poor. Here we describe the development of a method to explore the multiplicity of C. daubneyi infection and patterns of the parasite’s emergence and spread, based on Illumina MiSeq deep sequencing of meta barcoded amplicons of a fragment of the cytochrome c oxidase subunit I (mt-COX-1) locus. Our results show high levels of genetic diversity in 32 C. daubneyi populations derived from finished prime cattle consigned to slaughter from northern United Kingdom. The results are consistent with a single introduction of C. daubneyi infection to some of the farms where the cattle had been grazed during their lifetime and multiple introductions to most. The results illustrate the impact of high levels of animal movements in the United Kingdom, whereby multiple common mt-COX-1 haplotypes were identified in 26 populations in the absence of geographical clustering of clades. This has implications for the adaptability of environmental and intermediate host stages of the parasite to changing climatic and animal management conditions, or of parasitic stages to exposure to anthelmintic drugs; potentially allowing for greater pathogenicity, or the development of anthelmintic resistance, respectively.
The development of antibiotic resistance in bacteria is a major public health threat. Infection rates of resistant pathogens continue to rise against nearly all antimicrobials, which has led to development of different strategies to combat the antimicrobial resistance. In this review, we discuss how the newly popular CRISPR-cas system has been applied to combat antibiotic resistance in both extracellular and intracellular pathogens. We also review a recently developed method in which nano-size CRISPR complex was used without any phage to target the mecA gene. However, there is still challenge to practice these methods in field against emerging antimicrobial resistant pathogens.
More than 70 species of the Superfamily Paramphistomoidea, have been identified in ruminants in different parts of the world. Most are pathogenic, causing amphistomosis. Adult flukes within this family have a predilection for the forestomach (rumen) or bile duct of the liver, where they may cause epithelial damage. Identification of adult Paramphistomum, Calicophoron, Gastrothylax and Fischoederius at the species level based on morphology requires specialised expertise, whereas molecular genetic marker analysis is more precise and transferable. In the present study, we performed molecular characterisation of twenty seven adult flukes collected from the forestomachs of buffalo, cattle and goats in the Punjab province of Pakistan. PCR and sequencing of the ITS-2 rDNA region revealed a single haplotype in all cases. Phylogenetic comparison of P. epiclitum ITS2-rDNA sequences with those from other Paramphistomum, Calicophoron, Gastrothylax and Fischoederius species was performed to assess within and between species variation and validate the use of ITS-2 rDNA as a robust species-specific marker for P. epiclitum identification. This work provides a validated species-specific marker of P. epiclitum and the first report of this parasite species from Pakistan. The results of this study also have implications for the diagnosis and control of rumen flukes in the region and the need for accurate species identification to understand parasite distribution and population genetics. [Display omitted] •Molecular confirmation of P. epiclitum using rDNA ITS-2 and the significance of its phylogenetic relationship to other fluke.•Differentiation between Paramphistomum, Gastrothylax and Fischoederius as an aid to their identification in the rumen.•The present study has important implications for the diagnosis and control of rumen flukes.
The population genetics of nematode parasites are poorly understood with practical reference to the selection and spread of anthelmintic resistance mutations. Haemonchus species are important to study the nematode population genetics due to their clinical importance in ruminant livestock, and the availability of genomic resources. In the present study, it has been examined that Haemonchus contortus and Haemonchus placei populations from three buffalo and nine cattle hosts. Seventy-three individual adult worms of H. contortus and 148 of H. placei were analysed using a panel of seven microsatellite markers. The number of alleles per locus in H. contortus and H. placei indicated that all populations were polymorphic for the microsatellites used in the present study. Genetic diversity parameters included high levels of allelic richness and heterozygosity, indicating effective population sizes, high mutation rates and high transmission frequencies in the area. Genetic structure parameters revealed low genetic differentiation between and high levels of genetic variation within H. contortus and H. placei populations. Population dynamic analyses showed an absence of heterozygosity excess in both species, suggesting that there was no deviation from genetic drift equilibrium. Our results provide a proof of concept for better understanding of the consequences of specific control strategies, climatic change or management strategies on the population genetics of anthelmintic resistance alleles in Haemonchus spp. infecting co-managed buffalo and cattle.
Fasciola spp. are responsible for over 3 billion US dollars of production loss annually in livestock and cause widespread zoonotic disease. Nevertheless, understating of the emergence and spread of the trematode species is poor. The multiplicity of F. gigantica infection and its spread is potentially influenced by multiple factors, including the abundance of suitable intermediate hosts, climatic conditions favouring the completion of the parasite's lifecycle, and translocation of infected animals, or free-living parasite stages between regions. Here we describe the development of a ‘tremabiome’ metabarcoding sequencing method to explore the numbers of F. gigantica genotypes per infection and patterns of parasite spread, based on genetic characteristics of the mitochondrial NADH dehydrogenase 1 (mt-ND-1) locus. We collected F. gigantica from three abattoirs in the Punjab and Balochistan provinces of Pakistan, and our results show a high level of genetic diversity in 20 F. gigantica populations derived from small and large ruminants consigned to slaughter in both provinces. This implies that F. gigantica can reproduce in its definitive hosts through meiosis involving cross- and self-breeding, as described in the closely related species, Fasciola hepatica. The genetic diversity between the 20 populations derived from different locations also illustrates the impact of animal movements on gene flow. Our results demonstrate the predominance of single haplotypes, consistent with a single introduction of F. gigantica infection in 85% of the hosts from which the parasite populations were derived. This is consistent with clonal reproduction in the intermediate snail hosts. [Display omitted] •To confirm the species identity of recovered Fasciola spp.•To identify the presence of single or multiple genotypes per infection (multiplicity of infection)•Demonstrate the spread of F. gigantica mt-ND-1 haplotypes
An extended range of host susceptibility including camel has been evidenced for some of the important veterinary and public health pathogens, such as brucellosis, peste des petits ruminants (PPR) and bluetongue (BT). However, in disease endemic settings across many parts of the globe, most of the disease control interventions accounts for small and large ruminants, whereas unusual hosts and/or natural reservoirs, such as camels, remain neglected for disease control measures including routine vaccination. Such a policy drawback not only plays an important role in disease epizootiology particularly in settings where disease is endemic, but also serves an obstacle in disease control and subsequent eradication in future. With this background, using pre-validated ELISA and molecular assays [multiplex PCR, reverse transcriptase (RT)-PCR and real-time (rt)-PCR], we conducted a large-scale pathogen- and antibody-based surveillance for brucellosis, peste des petits ruminants and bluetongue in camel population (n = 992) originating from a wide geographical region in southern part of the Punjab province, Pakistan. Varying in each of the selected districts, the seroprevalence was found to be maximum for bluetongue [n = 697 (70.26%, 95% CI: 67.29–73.07)], followed by PPR [n = 193 (19.46%, 95% CI: 17.07–22.09)] and brucellosis [n = 66 (6.65%, 95% CI: 5.22–8.43)]. Odds of seroprevalence were more significantly associated with pregnancy status (non-pregnant, OR = 2.23, 95% CI: 1.86–5.63, p
Foot and mouth disease (FMD) is a viral disease that affects predominantly cloven-footed animal species within the order Artiodactyla. The potential of the virus to transmit, maintain and circulate itself across a wide range of susceptible hosts, including both domestic and wild ungulates, remains a single major obstacle in an effective eradication of disease worldwide, particularly in disease-endemic settings. Hence, a better understanding of virus transmission dynamics is very much crucial for an efficient control of the disease, particularly at places or regions where wildlife and livestock rearing co-exists. Both OIE and FAO have jointly launched the FMD-control program as FMD-Progressive Control Pathway (PCP) in various disease-endemic developing countries. Nevertheless, the propensity of virus to inter- and intra-species transmission may be a possible constraint in disease control and, hence, its subsequent eradication in such countries. Other than this, cross-species transmission, among domestic and wild ungulates living in close proximities, can undermine the conservation efforts for endangered species. We reviewed and summarized the so-far available information about inter- and intra-species disease transmission, and its impact on wildlife populations to better comprehend disease epidemiology and substantiate efforts for eventual disease eradication across the globe, particularly in settings where the disease is endemic.
Parasitic infestations are continuously posing enormous threats to food security and poverty alleviation. While different drugs are proposed to minimize the impact of parasite, these come with established genetic resistance. Understanding markers of genetic resistance will streamline the process of use of alternative medication as well as assessing the affect of currently applied anthelmintic. This review briefly outlines the roles of parasites in diseases and proposes the use of different genetic markers to mitigate the emerging resistance in parasite.
Newcastle disease (ND), caused by virulent 1 (AAvV 1), affects a wide range of avian species worldwide. Recently, several AAvVs of diverse genotypes have emerged with varying genomic and residue substitutions, and subsequent clinical impact on susceptible avian species. We assessed the clinico-pathological influence of two different AAvV 1 pathotypes [wild bird originated-velogenic strain (sub-genotype VIIi, MF437287) and feral pigeon originated-mesogenic strain (sub-genotype VIm, KU885949)] in commercial broiler chickens and pigeons. The velogenic strain caused 100% mortality in both avian species while the mesogenic strain caused 0% and 30% mortality in chickens and pigeons, respectively. Both strains showed tissue tropism for multiple tissues including visceral organs; however, minor variances were observed according to host and pathotype. The observed gross and microscopic lesions were typical of AAvV 1 infection. Utilizing oropharyngeal and cloacal swabs, a comparable pattern of viral shedding was observed for both strains from each of the infected individuals of both avian species. The study concludes a varying susceptibility of chickens and pigeons to different wild bird-originated AAvV 1 pathotypes and, therefore, suggests continuous monitoring and surveillance of currently prevailing strains for effective control of the disease worldwide, particularly in disease-endemic countries.
Since its first report in 1942, peste-des-petits-ruminants virus (PPRV) has caused several epidemics in a wide range of susceptible hosts around the world. In the last 30 years, the evidence of natural and experimental infections and virus isolation were reported from novel but unusual hosts such as camel, cattle, buffalo, dogs, Asiatic lion and pigs. In addition, PPRV in a potential vector, biting midges (Culicoides imicola), has been reported. Either presented as clinical and/or subclinical infections, the presence of the virus in an extended range of susceptible hosts highlights the cross-species transmission and supports the hypothesis of an endemic circulation of PPRV among susceptible hosts. However, the potential role of large ruminants, camels and unusual hosts for PPRV epidemiology is still obscure. Therefore, there is a need for molecular and epidemiological investigations of the disease among usual and unusual hosts to achieve the goals of disease control and eradication programmes initiated by national and international organisations, such as the FAO and OIE. This review is the first to summarise the scattered data on PPR in large ruminants, camels and unusual hosts to obtain the global scientific communities' attention for further research on epidemiological aspects, not only in its native hosts, but also in large ruminants, camels and other unusual hosts.
Pyrimethamine resistance is a major concern for the control of human haemoprotozoa, especially Plasmodium species. Currently, there is little understanding of how pyrimethamine resistance developed in Plasmodium vivax in the natural field conditions. Here, we present for the first time evidence of positive selection pressure on a dihydrofolate reductase locus and its consequences on the emergence and the spread of pyrimethamine resistance in P. vivax in the Punjab province of Pakistan. First, we examined the dihydrofolate reductase locus in 38 P. vivax isolates to look for evidence of positive selection pressure in human patients. The S58R (AGA)/S117N (AAC) double mutation was most common, being detected in 10/38 isolates. Single mutation S117N (AAC), I173L (CTT) and S58R (AGA) SNPs were detected in 8/38, 2/38 and 1/38 isolates, respectively. The F57L/I (TTA/ATA) and T61M (ATG) SNPs were not detected in any isolates examined. Although both soft and hard selective sweeps have occurred with striking differences between isolates, there was a predominance of hard sweeps. A single resistance haplotype was present at high frequency in 9/14 isolates, providing a strong evidence for single emergence of resistance by the single mutation, characteristics of hard selective sweeps. In contrast, 5/14 isolates carried multiple resistance haplotypes at high frequencies, providing an evidence of the emergence of resistance by recurrent mutations, characteristics of soft selective sweeps. Our phylogenetic relationship analysis suggests that S58R (AGA)/S117N (AAC) and S117N (AAC) mutations arose multiple times from a single origin and spread to multiple different cities in the Punjab province through gene flow. Interestingly, the I173L (CTT) mutation was present on a single haplotype, suggesting that it arises rarely and has not spread between cities. Our work shows the need for responsible use of existing and new antimicrobial drugs and their combinations, control the movement of infected patients and mosquito vector control strategies. •Illumina MiSeq deep amplicon sequencing was used to identify pyrimethamine resistance associated SNPs in Plasmodium vivax.•Pyrimethamine resistance was identified in field populations.•Positive selection pressure for resistance in human patients was investigated.•Hard and soft selective sweep models were used for the emergence of resistance.•Both single and multiple recurrent resistance mutations were spread by migration.
Gastrointestinal nematodes (GIN) are a serious concern for sheep producers worldwide. However, there is a paucity of evidence describing the epidemiology of GIN on modern UK sheep farms. The aim of this paper was to understand whether expected seasonal variations of infection are still found in ewes and lambs under varying management strategies in temperate climates. Faecal egg counts (FEC) were conducted on freshly voided samples collected from groups of ewes and lambs every third week for twelve months on three farms in southeast Scotland. The patterns of egg output have been described here in relation to management practices undertaken on the farms. Despite changes in farming practice and climatic conditions, the findings complement historical studies detailing the epidemiology of GIN. Findings include a periparturient rise in ewe FEC on two of the farms, while lambing time treatment appeared to suppress this on the third farm. On the same two farms lamb FEC increased during the summer, reaching a peak in the autumn. The work also highlights how the ad hoc use of anthelmintics does little to impact these patterns.
Anthelmintic resistance is a threat to global food security. In order to alleviate the selection pressure for resistance and maintain drug efficacy, management strategies increasingly aim to preserve a proportion of the parasite population in ‘refugia’, unexposed to treatment. While persuasive in its logic, and widely advocated as best practice, evidence for the ability of refugia-based approaches to slow the development of drug resistance in parasitic helminths is currently limited. Moreover, the conditions needed for refugia to work, or how transferable those are between parasite-host systems, are not known. This review, born of an international workshop, seeks to deconstruct the concept of refugia and examine its assumptions and applicability in different situations. We conclude that factors potentially important to refugia, such as the fitness cost of drug resistance, the degree of mixing between parasite sub-populations selected through treatment or not, and the impact of parasite life-history, genetics and environment on the population dynamics of resistance, vary widely between systems. The success of attempts to generate refugia to limit anthelmintic drug resistance are therefore likely to be highly dependent on the system in hand. Additional research is needed on the concept of refugia and the underlying principles for its application across systems, as well as empirical studies within systems that prove and optimise its usefulness. [Display omitted] •Refugia is currently recommended for strategic control of AR in GI nematodes of livestock.•Conditions required for refugia-based control may differ between parasite systems.•An improved understanding of the parameters influencing refugia-based control is required.
A study was designed to improve understanding of the genetics of Theileria annulata populations in sympatric cattle and Asian buffalo (Bubalus bubalus). The study was undertaken in the Punjab province of Pakistan, where the prevalence of tropical theileriosis is high. Parasite materials were collected from infected animals in defined regions, where cattle and Asian buffalo are kept together. Six satellite DNA markers and a mitochondrial cytochrome b marker were used to explore the multiplicity of T. annulata infection and patterns of emergence and spread of different parasite genotypes. The results show differences in the numbers of unique satellite locus alleles, suggesting that T. annulata is genetically more diverse in cattle- than in buffalo-derived populations. Heterozygosity (He) indices based on satellite and cytochrome b loci data show high levels of genetic diversity among the cattle- and buffalo-derived T. annulata populations. When considered in the context of high parasite transmission rates and frequent animal movements between different regions, the predominance of multiple T. annulata genotypes and multiple introductions of infection may have practical implications for the spread of parasite genetic adaptations; such as those conferring vaccine cross-protection against different strains affecting cattle and Asian buffalo, or resistance to antiprotozoal drugs.
With extensive use of the anthelmintic in many tropical and parasite-endemic countries, the graveness of parasite resistance is exaggerating. The trend in the use of excessive medication in animal is also posing threats to public health. The governmental bodies are either lacking understanding or underestimating the regime of this emergingissue. This brief overview highlights the need, rationale and urgency of the matter to establish foundations for unifiedaction plans at the national levels as well as international levels in mitigating the development of resistance.
Internal parasites are a major concern in livestock production because they can impact the health and well-being of animals clinically and subclinically, and ultimately cause significant production loss. Among these internal parasites are nematodes, tapeworms, flukes, and coccidian protozoans. This review focuses on the diagnostic tests that are routinely performed by veterinarians and diagnostic laboratories, but also highlights recently developed tools that may improve diagnostic capabilities, including molecular and immunodiagnostic tests. Overall, diagnostic tests for parasites of livestock are an integral part of health management practices, and for assessing individual animal and herd health.
•F200Y (TAC) benzimidazole resistance mutation in H. placei was found on a single haplotype in multiple populations across Punjab province.•F200Y (TAC) mutations was found on multiple divergent haplotypes in H. contortus.•The P167 (TAC) and E198A (GCA) mutations were not detected in any H. contortus and H. placei populations. Benzimidazoles have been intensively (for over 40 years) used in the livestock sector, particularly in small ruminants. This has been led to the widespread emergence of resistance in a number of small ruminant parasite species, especially Haemonchus contortus. In many countries benzimidazole resistance has severely compromised the control of H. contortus in small ruminants; but there is a little information on benzimidazole resistance in H. contortus infecting buffalo and cattle. Resistance to benzimidazoles have also been reported in the large ruminant parasite, Haemonchus placei, but again there is relatively little information on its prevalence. Hence it is very important to understand how resistance-conferring mutations emerge and spread in both parasites in buffalo and cattle hosts in order to develop approaches for the recognition of the problem at an early stage of its development. The present study suggests that the F200Y (TAC) mutation is common in H. contortus, being detected in 5/7 populations at frequencies between 7 and 57%. Furthermore, 6/10 H. placei populations contained the F200Y (TAC) mutation, albeit at low frequencies of between 0.4 and 5%. The phylogenetic analysis suggests that the F200Y (TAC) mutation in H. contortus has emerged on multiple occasions in the region, with at least three independent emergences across the populations. In contrast, the F200Y (TAC) resistance-conferring mutation in H. placei is only seen on a single haplotype. A high level frequency of the resistance haplotypes in the region, suggests that the unique resistance conferring-mutation has spread from a single emergence; likely by anthropogenic animal movement. Overall, these results provide the first clear genetic evidence for the spread of benzimidazole resistance-conferring mutations to multiple different locations from a single emergence in H. placei; while being consistent with previous small ruminant-based observations of multiple emergence of resistance mutations in H. contortus.
Varestrongylus eleguneniensis (Nematoda; Protostrongylidae) is a recently described species of lungworm that infects caribou (Rangifer tarandus), muskoxen (Ovibos moschatus) and moose (Alces americanus) across northern North America. Herein we explore the geographic distribution of V. eleguneniensis through geographically extensive sampling and discuss the biogeography of this multi-host parasite. We analyzed fecal samples of three caribou subspecies (n = 1485), two muskox subspecies (n = 159), and two moose subspecies (n = 264) from across northern North America. Protostrongylid dorsal-spined larvae (DSL) were found in 23.8%, 73.6%, and 4.2% of these ungulates, respectively. A portion of recovered DSL were identified by genetic analyses of the ITS-2 region of the nuclear rDNA or the cytochrome oxidase c subunit I (COI) region of the mtDNA. We found V. eleguneniensis widely distributed among caribou and muskox populations across most of their geographic prange in North America but it was rare in moose. Parelaphostrongylus andersoni was present in caribou and moose and we provide new geographic records for this species. This study provides a substantial expansion of the knowledge defining the current distribution and biogeography of protostrongylid nematodes in northern ungulates. Insights about the host and geographic range of V. eleguneniensis can serve as a geographically extensive baseline for monitoring current distribution and in anticipating future biogeographic scenarios under a regime of accelerating climate and anthropogenic perturbation. [Display omitted] •Varestrongylus eleguneniensis is a lungworm whose primary host is the caribou.•The muscleworm, Parelaphostrongylus andersoni, co-infects caribou across its range.•We expand the knowledge on distribution of the caribou lungworm and the muscleworm.•Muskoxen sympatric with caribou are infected with the caribou lungworm.•We discuss the biogeography of V. eleguneniensis and Rangifer across North America.
Piroplasmosis is caused by tick-borne haemoprotozoa of the genera Theileria and Babesia. These parasitic infections can seriously impact on the health of livestock and production. Piroplasms of multiple species can be present in a single host, but reliable molecular diagnostic tools are needed in order to understand the composition of these complex parasite communities. Theileria and Babesia vary in their epidemiology, drug sensitivity, pathogenicity and interaction with co-infecting species, but are similar in that infected animals become persistent carriers after recovery from primary infection, acting as reservoir hosts. Here, we describe for the first time the use of a deep amplicon sequencing platform to identify proportions of piroplasm species in co-infecting communities and develop the concept of a “haemoprotobiome”. First, four phenotypically-verified species of Theileria and Babesia were used to prepare mock DNA pools with random numbers of the parasites amplified by four different numbers of PCR cycles to assess sequence representation for each species. Second, we evaluated the detection threshold of the deep amplicon sequencing assay for each of the four species and to assess the accuracy of proportional quantification of all four species. Finally, we applied the assay to the field samples to afford insight of the species composition of piroplasm communities in small and large ruminants in the Punjab province of Pakistan. The “haemoprotobiome” concept has several potential applications in veterinary and human research, including understanding of responses to drug treatment; parasite epidemiology and ecology; species interactions during mixed infections; and parasite control strategies.
Coronaviruses (CoVs) are enveloped, positive sense, single-stranded RNA viruses. The viruses have adapted to infect a large number of animal species, ranging from bats to camels. At present, seven CoVs infect humans, of which Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) is responsible for causing the Coronavirus Disease 2019 (COVID-19) in humans. Since its emergence in late 2019, SARS-CoV-2 has spread rapidly across the globe. Healthcare systems around the globe have been stretched beyond their limits posing new challenges to emergency healthcare services and critical care. The outbreak continues to jeopardize human health, social life and economy. All known human CoVs have zoonotic origins. Recent detection of SARS-CoV-2 in pet, zoo and certain farm animals has highlighted its potential for reverse zoonosis. This scenario is particularly alarming, since these animals could be potential reservoirs for secondary zoonotic infections. In this article, we highlight interspecies SARS-CoV-2 infections and focus on the reverse zoonotic potential of this virus. We also emphasize the importance of potential secondary zoonotic events and the One-Health and One-World approach to tackle such future pandemics.
Lancet liver flukes of the genus Dicrocoelium (Trematoda: Digenea) are recognised parasites of domestic and wild herbivores. The aim of the present study was to confirm the species identity of Dicrocoeliid flukes collected from the Chitral valley in the Himalayan ranges of Pakistan. The morphology of 48 flukes belonging to eight host populations was examined; but overlapping traits prevented accurate species designation. Phylogenetic comparison of published D. dendriticum ribosomal cistron DNA, and cytochrome oxidase-1 (COX-1) mitochondrial DNA sequences with those from D. chinensis was performed to assess within and between species variation and re-affirm the use of species-specific single nucleotide polymorphism markers. PCR and sequencing of 34 corresponding fragments of ribosomal DNA and 14 corresponding fragments of mitochondrial DNA from the Chitral valley flukes, revealed 10 and 4 unique haplotypes, respectively. These confirmed for the first time the molecular species identity of Pakistani lancet liver flukes as D. dendriticum. This work provides a preliminary illustration of a phylogenetic approach that could be developed to study the ecology, biological diversity, and epidemiology of Dicrocoeliid lancet flukes when they are identified in new settings. •First molecular confirmation of Dicrocoelium dendriticum in Himalayan Pakistan.•Use of ribosomal and mitochondrial DNA phylogenetic markers.•Demonstration of the complementary value of morphological and molecular speciation methods for Dicrocoeliid flukes.
Parasitic gastrointestinal nematodes contribute to significant human morbidity and cause billions of dollars per year in lost agricultural production. Control is dependent on the use of anthelmintic drugs which, in the case of livestock parasites, is severely compromised by the widespread development of drug resistance. There are now concerns regarding the emergence of anthelmintic resistance in parasitic nematodes of humans in response to the selection pressure resulting from mass drug administration programs. Consequently, there is an urgent need for sensitive, scalable and accurate diagnostic tools to detect the emergence of anthelmintic resistance. Detecting and measuring the frequency of resistance-associated mutations in parasite populations has the potential to provide sensitive and quantitative assessment of resistance emergence from an early stage. Here, we describe the development and validation of deep amplicon sequencing as a powerful new approach to detect and quantify the frequency of single nucleotide polymorphisms associated with benzimidazole resistance. We have used parasite communities in sheep to undertake a proof-of-concept study of this approach. Sheep provide an excellent host system, as there are multiple co-infecting trichostrongylid nematode species, each likely with a varying prevalence of benzimidazole resistance. We demonstrate that the approach provides an accurate measure of resistance allele frequencies, and can reliably detect resistance alleles down to a frequency of 0.1%, making it particularly valuable for screening mutations in the early stages of resistance. We illustrate the power of the technique by screening UK sheep flocks for benzimidazole resistance-associated single nucleotide polymorphisms at three different codons of the β-tubulin gene in seven different parasite species from 164 populations (95 from ewes and 69 from lambs) in a single MiSeq sequencing run. This approach provides a powerful new tool to screen for the emergence of anthelmintic resistance mutations in parasitic nematode populations of both animals and humans.
A recurrent mite infestation affecting a room used to inspect fabric in a UK textile mill was investigated to allay concerns of any potential health risks to factory staff, and to inform the unknown risk of downgrading of the product. The approach integrated conventional morphological examination of adult female mites by referring to published identification keys, with molecular speciation based on amplification of a 16S ribosomal DNA fragment. The methods enabled the mites to be unambiguously identified as Dermanyssus gallinae ‘special lineage L1’. Subsequent investigations showed the source of infestation to be pigeons nesting in the air ducts, with the gamasid mites moving into the room once the young birds had fledged. This is the first report of D. gallinae ‘special lineage L1’ in northern Europe. Previous reports of nosocominal gamasoidosis caused by D. gallinae ‘special lineage L1’ originating from feral pigeon populations have been from southern Europe. Confirmation of the mite identity was important in allowing the mill to take remedial and preventive action. In this clinical communication, we provide images of the key morphological features used to identify D. gallinae and describe a molecular protocol to confirm ‘special lineage L1’. •Investigation of mite infestation in a textile factory•Morphological keys to identify poultry red mites•Molecular speciation of Dermanyssus gallinae and confirmation of ‘special lineage L1’•First report of D. gallinae ‘special lineage L1’ in northern Europe•Risk of zoonotic gamasoidosis from pigeons nesting in air ducts
Haemonchus contortus is a globally distributed and economically important gastrointestinal pathogen of small ruminants and has become a key nematode model for studying anthelmintic resistance and other parasite-specific traits among a wider group of parasites including major human pathogens. Here, we report using PacBio long-read and OpGen and 10X Genomics long-molecule methods to generate a highly contiguous 283.4 Mbp chromosome-scale genome assembly including a resolved sex chromosome for the MHco3(ISE).N1 isolate. We show a remarkable pattern of conservation of chromosome content with Caenorhabditis elegans, but almost no conservation of gene order. Short and long-read transcriptome sequencing allowed us to define coordinated transcriptional regulation throughout the parasite’s life cycle and refine our understanding of cis- and trans-splicing. Finally, we provide a comprehensive picture of chromosome-wide genetic diversity both within a single isolate and globally. These data provide a high-quality comparison for understanding the evolution and genomics of Caenorhabditis and other nematodes and extend the experimental tractability of this model parasitic nematode in understanding helminth biology, drug discovery and vaccine development, as well as important adaptive traits such as drug resistance.
Various PCR based methods have been described for the diagnosis of malaria, but most depend on the use of Plasmodium species-specific probes and primers; hence only the tested species are identified and there is limited available data on the true circulating species diversity. Sensitive diagnostic tools and platforms for their use are needed to detect Plasmodium species in both clinical cases and asymptomatic infections that contribute to disease transmission. We have recently developed for the first time a novel high throughput ‘haemoprotobiome’ metabarcoded DNA sequencing method and applied it for the quantification of haemoprotozoan parasites (Theleria and Babesia) of livestock. Here, we describe a novel, high throughput method using an Illumina MiSeq platform to demonstrate the proportions of Plasmodium species in metabarcoded DNA samples derived from human malaria patients. Plasmodium falciparum and Plasmodium vivax positive control gDNA was used to prepare mock DNA pools of parasites to evaluate the detection threshold of the assay for each of the two species. The different mock pools demonstrate the accurate detection ability and to show the proportions of each of the species being present. We then applied the assay to malaria-positive human samples to show the species composition of Plasmodium communities in the Punjab province of Pakistan and in the Afghanistan-Pakistan tribal areas. The diagnostic performance of the deep amplicon sequencing method was compared to an immunochromatographic assay that is widely used in the region. The deep amplicon sequencing showed that P. vivax was present in 69.8%, P. falciparum in 29.5% and mixed infection in 0.7% patients examined. The immunochromatographic assay showed that P. vivax was present in 65.6%, P. falciparum in 27.4%, mixed infection 0.7% patients and 6.32% malaria-positive cases were negative in immunochromatographic assay, but positive in the deep amplicon sequencing. Overall, metabarcoded DNA sequencing demonstrates better diagnostic performance, greatly increasing the estimated prevalence of Plasmodium infection. The next-generation sequencing method using metabarcoded DNA has potential applications in the diagnosis, surveillance, treatment, and control of Plasmodium infections, as well as to study the parasite biology. •We reported for the first time the development of Haemoprotobiome technology to quantify the P. falciparum and P. vivax.•P. falciparum and P. vivax mock pools demonstrate the accurate detection ability and to show the proportions of each of the species.•Haemoprotobiome demonstrates better diagnostic performance than immunochromatographic assay.
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