| 1. |
- Johansson, Håkan, 1972-, et al.
(författare)
-
Characterization of Campylobacter spp. isolated from wild birds in the Antarctic and Sub-Antarctic
- 2018
-
Ingår i: PLOS ONE. - : Public Library of Science. - 1932-6203. ; 13:11
-
Tidskriftsartikel (refereegranskat)abstract
- A lack of knowledge of naturally occurring pathogens is limiting our ability to use the Antarctic to study the impact human-mediated introduction of infectious microorganisms have on this relatively uncontaminated environment. As no large-scale coordinated effort to remedy this lack of knowledge has taken place, we rely on smaller targeted efforts to both study present microorganisms and monitor the environment for introductions. In one such effort, we isolated Campylobacter species from fecal samples collected from wild birds in the Antarctic Peninsula and the sub-Antarctic island of South Georgia. Indeed, in South Georgia, we found Campylobacter lari and the closely related Campylobacter peloridis, but also distantly related human-associated multilocus sequence types of Campylobacter jejuni. In contrast, in the Antarctic Peninsula, we found C. tart and two closely related species, Campylobacter subantarcticus and Campylobacter volucris, but no signs of human introduction. In fact, our finding of human-associated sequence types of C. jejuni in South Georgia, but not in the Antarctic Peninsula, suggests that efforts to limit the spread of infectious microorganisms to the Antarctic have so far been successful in preventing the introduction of C. jejuni. However, we do not know how it came to South Georgia and whether the same mode of introduction could spread it from there to the Antarctic Peninsula.
|
|
| 2. |
- Ahlinder, J., et al.
(författare)
-
Use of metagenomic microbial source tracking to investigate the source of a foodborne outbreak of cryptosporidiosis
- 2022
-
Ingår i: FOOD AND WATERBORNE PARASITOLOGY. - : Elsevier. - 2405-6766. ; 26
-
Tidskriftsartikel (refereegranskat)abstract
- Cryptosporidium is a protozoan parasite of global public health importance that causes gastroenteritis in a variety of vertebrate hosts, with many human outbreaks reported yearly, often from ingestion of contaminated water or food. Despite the major public health implications, little is typically known about sources of contamination of disease outbreaks caused by Cryptosporidium. Here, we study a national foodborne outbreak resulted from infection with Cryptosporidium parvum via romaine lettuce, with the main goal to trace the source of the parasite. To do so, we combined traditional outbreak investigation methods with molecular detection and characterization methods (i.e. PCR based typing, amplicon and shotgun sequencing) of romaine lettuce samples collected at the same farm from which the contaminated food was produced. Using 18S rRNA typing, we detected C. parvum in two out of three lettuce samples, which was supported by detections in the metagenome analysis. Microbial source tracking analysis of the lettuce samples suggested sewage water as a likely source of the contamination, albeit with some uncertainty. In addition, the high degree of overlap in bacterial species content with a public human gut microbial database corroborated the source tracking results. The combination of traditional and molecular based methods applied here is a promising tool for future source tracking investigations of food- and waterborne outbreaks of Cryptosporidium spp. and can help to control and mitigate contamination risks.
|
|
| 3. |
- Alsmark, UCM, et al.
(författare)
-
Two outbreaks of cryptosporidiosis associated with cattle spring pasture events
- 2018
-
Ingår i: Veterinary Parasitology. - : Elsevier BV. - 2405-9390. ; 14, s. 71-74
-
Tidskriftsartikel (refereegranskat)abstract
- Over a period of less than four weeks, 50 human cases of cryptosporidiosis were reported from a relatively small geographical area in Sweden. All cases were associated with visits to cattle spring pasture events at two farms (referred to as Farm A and B). Epidemiological and microbiological evidence show that contact with calves at the farms was the most likely source of Cryptosporidium infections. Gp60 sequences from human and calf isolates at Farm A were identical to each other, but differed from those at Farm B where, again, human and calf gp60 sequences were identical, proving that the two outbreaks had no common origin. As a direct consequence of these two outbreaks, and guided by knowledge gained from the outbreak investigations, the Swedish Board of Agriculture and all relevant farmer advisory organizations have updated their hygiene instructions for farm visits.
|
|
| 4. |
- Ankarklev, Johan, et al.
(författare)
-
A novel high-resolution multilocus sequence typing of Giardia intestinalis Assemblage A isolates reveals zoonotic transmission, clonal outbreaks and recombination
- 2018
-
Ingår i: Infection, Genetics and Evolution. - : Elsevier BV. - 1567-1348 .- 1567-7257. ; 60, s. 7-16
-
Tidskriftsartikel (refereegranskat)abstract
- Molecular epidemiology and genotyping studies of the parasitic protozoan Giardia intestinalis have proven difficult due to multiple factors, such as low discriminatory power in the commonly used genotyping loci, which has hampered molecular analyses of outbreak sources, zoonotic transmission and virulence types. Here we have focused on assemblage A Giardia and developed a high-resolution assemblage-specific multilocus sequence typing (MLST) method. Analyses of sequenced G. intestinalis assemblage A genomes from different sub-assemblages identified a set of six genetic loci with high genetic variability. DNA samples from both humans (n = 44) and animals (n = 18) that harbored Giardia assemblage A infections, were PCR amplified (557-700 bp products) and sequenced at the six novel genetic loci. Bioinformatic analyses showed five to ten-fold higher levels of polymorphic sites than what was previously found among assemblage A samples using the classic genotyping loci. Phylogenetically, a division of two major clusters in assemblage A became apparent, separating samples of human and animal origin. A subset of human samples (n = 9) from a documented Giardia outbreak in a Swedish day-care center, showed full complementarity at nine genetic loci (the six new and the standard BG, TPI and GDH loci), strongly suggesting one source of infection. Furthermore, three samples of human origin displayed MLST profiles that were phylogenetically more closely related to MLST profiles from animal derived samples, suggesting zoonotic transmission. These new genotyping loci enabled us to detect events of recombination between different assemblage A isolates but also between assemblage A and E isolates. In summary, we present a novel and expanded MLST strategy with significantly improved sensitivity for molecular analyses of virulence types, zoonotic potential and source tracking for assemblage A Giardia.
|
|
| 5. |
- Ankarklev, Johan, et al.
(författare)
-
Behind the smile : cell biology and disease mechanisms of Giardia species
- 2010
-
Ingår i: Nature Reviews Microbiology. - : Springer Science and Business Media LLC. - 1740-1526 .- 1740-1534. ; 8:6, s. 413-422
-
Forskningsöversikt (refereegranskat)abstract
- The eukaryotic intestinal parasite Giardia intestinalis was first described in 1681, when Antonie van Leeuwenhoek undertook a microscopic examination of his own diarrhoeal stool. Nowadays, although G. intestinalis is recognized as a major worldwide contributor to diarrhoeal disease in humans and other mammals, the disease mechanisms are still poorly understood. Owing to its reduced complexity and proposed early evolutionary divergence, G. intestinalis is used as a model eukaryotic system for studying many basic cellular processes. In this Review we discuss recent discoveries in the molecular cell biology and pathogenesis of G. intestinalis.
|
|
| 6. |
|
|
| 7. |
- Björkman, Camilla, et al.
(författare)
-
Cryptosporidium infections in suckler herd beef calves
- 2015
-
Ingår i: Parasitology. - 0031-1820 .- 1469-8161. ; 142, s. 1108-1114
-
Tidskriftsartikel (refereegranskat)abstract
- A study was carried out to investigate how common Cryptosporidium infections are in beef calves in Swedish suckler herds and to explore which species and subtypes that occur. We further aimed at identifying factors associated with shedding of Cryptosporidium oocysts in this type of calf management. The study was conducted in two regions in Sweden and included 30 herds. Faecal samples were collected from calves younger than 3 months. A brief clinical examination was done and a questionnaire was used to collect data on management routines. Faeces were cleaned and concentrated and oocysts identified by epifuorescence microscopy. Cryptosporidium positive samples were analyzed at the 18S rRNA and GP60 genes to determine species and Cryptosporidium parvum subtype, respectively. Logistic regression was used to identify factors associated with infection. Oocysts were detected in 122 (36·7%) calves from 29 (97%) herds, at 400 to 2·4 × 107 OPG. The youngest positive calves were only 1 and 2 days old. There was no association between age and Cryptosporidium infection. Cryptosporidium bovis, C. ryanae, C. parvum and C. ubiquitum were identified, with C. bovis being the major species. Two C. parvum subtypes, IIaA16G1R1 and IIdA27G1 were identified. Routines for cleaning calf pens and number of cows in calving pens were associated with infection.
|
|
| 8. |
- Björkman, Camilla, et al.
(författare)
-
Disinfection with hydrated lime may help manage cryptosporidiosis in calves
- 2018
-
Ingår i: Veterinary Parasitology. - : Elsevier BV. - 0304-4017 .- 1873-2550. ; 264, s. 58-63
-
Tidskriftsartikel (refereegranskat)abstract
- Diarrhea is common in young calves and is often caused by Cryptosporidium parvum infection. The aim of this study was to investigate if disinfection of calf pens with hydrated lime would reduce contamination of C. parvum oocysts and improve calf health in herds with C. parvum associated diarrhea problems. Four dairy herds with ongoing C. parvum associated calf diarrhea problems each participated in the study over six to seven months. During the study period, all pens/huts for young calves were cleaned according to the usual farm routine before a new calf entered. Hydrated lime was then used to disinfect half of the pens/huts. Diarrhea incidence was recorded by the farmers and by veterinarians, who clinically examined the calves every second month. In total, 402 calves participated in the study. The farmers detected diarrhea in 214 (53%) calves, with similar proportions in calves kept in lime disinfected and control pens. Age at diarrhea onset was significantly higher in lime disinfected pens than in control pens, 9.0 days and 7.6 days, respectively. There was no difference between the groups regarding duration or severity index recorded by the farmers. The body condition score in 6-8 week old calves was significantly higher in calves that had been kept in lime disinfected pens during their first weeks of life, indicating that calves in disinfected pens/huts were less affected by their infections.Faecal samples from 5 to 21 day old calves, were collected on four occasions at each farm (n = 95). Cryptosporidium positive samples were found at all samplings in all four herds. Cryptosporidium spp. was detected in 79 (83%) samples with no difference between lime disinfected and control pens. C. parvum was the dominant species. Two different C. parvum subtypes were found; IIaA16G1R1b in three herds and IIaA16G1R1b_variant in one herd. Only one subtype was found in each herd.Disinfection of calf pens with slaked lime delayed onset of diarrhea and improved the body condition in the calves, but did not affect diarrhea incidence or duration. Although lime disinfection alone will not be sufficient to control Cryptosporidium associated diarrhea in herds with extensive calf diarrhea problems, these results suggest that it can be a valuable complement to other measures.
|
|
| 9. |
- Bujila, Ioana, et al.
(författare)
-
Cryptosporidium chipmunk genotype I : An emerging cause of human cryptosporidiosis in Sweden
- 2021
-
Ingår i: Infection, Genetics and Evolution. - : Elsevier. - 1567-1348 .- 1567-7257. ; 92
-
Tidskriftsartikel (refereegranskat)abstract
- Most cases of cryptosporidiosis in humans are caused by Cryptosporidium parvum or Cryptosporidium hominis. However, more uncommon species are increasingly being recognised to cause infection in humans. Here we report that Cryptosporidium chipmunk genotype I, which has various rodents as its natural host, is the third most common source of human cryptosporidiosis in Sweden. We also describe the first small outbreak of cryptosporidiosis caused by Cryptosporidium chipmunk genotype I and report the first case of zoonotic transmission of Cryptosporidium chipmunk genotype I from a red squirrel to a human. Cryptosporidium chipmunk genotype I was identified in 20 human cases, including 16 sporadic cases, three outbreak-related cases, and one zoonotic case, as well as in two squirrel samples. Gp60 subtyping which was successful for 19 human cases and two squirrel samples showed that all samples harboured the same subtype, XIVaA20G2T1. The work presented here suggests that red squirrel is a natural host of Cryptosporidium chipmunk genotype I and that infection with Cryptosporidium chipmunk genotype I is an emerging cause of domestic cryptosporidiosis in Sweden and a potential source of outbreaks.
|
|
| 10. |
- Bujila, Ioana, et al.
(författare)
-
Cryptosporidium species and subtypes identified in human domestic cases through the national microbiological surveillance programme in Sweden from 2018 to 2022
- 2024
-
Ingår i: BMC Infectious Diseases. - : BMC. - 1471-2334. ; 24:1
-
Tidskriftsartikel (refereegranskat)abstract
- Background The intestinal protozoan parasite Cryptosporidium is an important cause of diarrheal disease worldwide. A national microbiological surveillance programme was implemented in Sweden in 2018 in order to increase knowledge of the molecular epidemiology of human cryptosporidiosis to better understand transmission patterns and potential zoonotic sources. This article summarises the results of the first five years of the surveillance programme. Methods Cryptosporidium-positive faecal and DNA samples from domestically acquired infections were collected from clinical microbiological laboratories in Sweden. Species and subtype determination was performed using 60 kDa glycoprotein and/or small subunit ribosomal RNA gene analysis. Results Between 2018 and 2022, 1654 samples were analysed and 11 different species were identified: C. parvum (n = 1412), C. mortiferum (n = 59), C. hominis (n = 56), C. erinacei (n = 11), C. cuniculus (n = 5), C. meleagridis (n = 3), C. equi (n = 2), C. ubiquitum (n = 2), and one each of C. canis, C. ditrichi and C. felis. Subtyping revealed seven subtype families of C. parvum (new subtype families IIy and IIz) and 69 different subtypes (11 new subtypes). The most common C. parvum subtypes were IIdA22G1c, IIdA24G1, IIdA15G2R1 and IIaA16G1R1b. For C. hominis, four different subtype families and nine different subtypes (two new subtypes) were identified. For additional species, two new subtype families (IIIk and VId) and nine new subtypes were identified. All successfully subtyped C. mortiferum cases were subtype XIVaA20G2T1, confirming previous findings in Sweden. Several outbreaks were identified of which the majority were foodborne and a few were due to direct contact with infected animals. Conclusion Infection with C. parvum is the leading cause of human cryptosporidiosis acquired in Sweden, where more than 90% of domestic cases are caused by this zoonotic species and only a small proportion of cases are due to infection with other species. The rodent-associated C. mortiferum is considered an emerging zoonotic species in Sweden and the number of domestically acquired human cases has surpassed that of infection with C. hominis. A high diversity of species and subtypes, as well as diversity within the same subtype, was detected. Also, cryptosporidiosis appears to affect adults to a great extent in Sweden.
|
|
| 11. |
- Deksne, Gunita, et al.
(författare)
-
Prevalence, risk factor and diversity of Cryptosporidium in cattle in Latvia
- 2022
-
Ingår i: Veterinary Parasitology. - : Elsevier. - 2405-9390. ; 28
-
Tidskriftsartikel (refereegranskat)abstract
- The epidemiology of Cryptosporidium spp. in Latvia was investigated by testing fecal samples from 926 animals aged from one day to 24 years for the presence of Cryptosporidium spp. oocysts. The samples were collected from 87 cattle farms and from four slaughterhouses, and analyzed by conventional and fluorescent microscopy, fol-lowed by Cryptosporidium species and C. parvum subtype differentiation. Moreover, using a questionnaire, we surveyed factors that could be relevant as risk factors of Cryptosporidium spp. infection on the farms. Crypto-sporidium spp. were shed by 33.8% of the investigated cattle and at least one shedding animal was found on 77.8% of the farms. In the present study, all four Cryptosporidium species reported to commonly infect cattle and two additional Cryptosporidium species (C. scrofarum and C. ubiquitum) were identified. In addition, mix in-fections of C. parvum/C. bovis, C. bovis/C. ryanae, C. parvum/C. ryanae, C. parvum/C. andersoni and C. bovis/ C. andersoni were observed. C. parvum and C. bovis was mostly prevalent in young animals (0-3 months old) and in addition, diarrhea associated with C. parvum infection was observed only in very young animals. Cryptosporidium andersoni and C. ryanae in age group 0-3 months was observed in low prevalence, while a higher proportion of animals with diarrhea associated with C. andersoni infection was observed in very young animals and with C. ryanae in animals age group 4-24 months. Eight previously described C. parvum subtypes were observed. The majority of the subtypes were in the IIa subtype family, while one subtype was identified from the IId subtype family. The most common subtype was IIaA15G2R1, which was found in 34.2% of the C. parvum successfully subtyped samples. The probability of Cryptosporidium spp. associated diarrhea in cattle decreased significantly with the age of the animals and a prolonged period during which calves were fed with milk.
|
|
| 12. |
- Dettwiler, Ines, et al.
(författare)
-
TIDE Analysis of Cryptosporidium Infections by gp60 Typing Reveals Obscured Mixed Infections
- 2022
-
Ingår i: Journal of Infectious Diseases. - : Oxford University Press (OUP). - 0022-1899 .- 1537-6613. ; 225:4, s. 686-695
-
Tidskriftsartikel (refereegranskat)abstract
- Background. Cryptosporidiosis is a parasitic disease associated with potentially fatal diarrhea. The most used method in Cryptosporidium subtyping is based on the glycoprotein gene gp60. Each infection can represent a parasite population, and it is important to investigate the influence on transmission and virulence, as well as any impact on public health investigations. However, an easy-to-use method for detection is lacking. Methods. Here we report on the use of the bioinformatic program TIDE for deconvolution of gp60 chromatograms. A combination of single oocyst analysis and cloning successfully confirmed the within-sample parasite population diversity. Retrospective sample analysis was conducted on archived chromatograms. Results. For Cryptosporidium parvum, 8.6% multistrain infections (13 of 152) obscured by currently used consensus base calling were detected. Importantly, we show that single oocysts can harbor a mixed population of sporozoites. We also identified a striking dominance of unappreciated polymerase stutter artefacts in all 218 chromatograms analyzed, challenging the uncritical use of gp60 typing. Conclusions. We demonstrate the value of a new, easy-to-use analytical procedure for critical characterization of C. parvum and Cryptosporidium hominis in epidemiological investigations, also applicable retrospectively. Our findings illuminate the hidden parasite diversity with important implications for tracing zoonotic and person-to-person transmissions.
|
|
| 13. |
- Deutsch, Lisa, 1965-
(författare)
-
Global trade, food production and ecosystem support : Making the interactions visible
- 2004
-
Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Modern food production is a complex, globalized system in which what we eat and how it is produced are increasingly disconnected. This thesis examines some of the ways in which global trade has changed the mix of inputs to food and feed, and how this affects food security and our perceptions of sustainability. One useful indicator of the ecological impact of trade in food and feed products is the Appropriated Ecosystem Areas (ArEAs), which estimates the terrestrial and aquatic areas needed to produce all the inputs to particular products.The method is introduced in Paper I and used to calculate and track changes in imported subsidies to Swedish agriculture over the period 1962-1994. In 1994, Swedish consumers needed agricultural areas outside their national borders to satisfy more than a third of their food consumption needs. The method is then applied to Swedish meat production in Paper II to show that the term “Made in Sweden” is often a misnomer. In 1999, almost 80% of manufactured feed for Swedish pigs, cattle and chickens was dependent on imported inputs, mainly from Europe, Southeast Asia and South America. Paper III examines ecosystem subsidies to intensive aquaculture in two nations: shrimp production in Thailand and salmon production in Norway. In both countries, aquaculture was shown to rely increasingly on imported subsidies. The rapid expansion of aquaculture turned these countries from fishmeal net exporters to fishmeal net importers, increasingly using inputs from the Southeastern Pacific Ocean.As the examined agricultural and aquacultural production systems became globalized, levels of dependence on other nations’ ecosystems, the number of external supply sources, and the distance to these sources steadily increased. Dependence on other nations is not problematic, as long as we are able to acknowledge these links and sustainably manage resources both at home and abroad. However, ecosystem subsidies are seldom recognized or made explicit in national policy or economic accounts. Economic systems are generally not designed to receive feedbacks when the status of remote ecosystems changes, much less to respond in an ecologically sensitive manner. Papers IV and V discuss the problem of “masking” of the true environmental costs of production for trade. One of our conclusions is that, while the ArEAs approach is a useful tool for illuminating environmentally-based subsidies in the policy arena, it does not reflect all of the costs. Current agricultural and aquacultural production methods have generated substantial increases in production levels, but if policy continues to support the focus on yield and production increases alone, taking the work of ecosystems for granted, vulnerability can result. Thus, a challenge is to develop a set of complementary tools that can be used in economic accounting at national and international scales that address ecosystem support and performance.We conclude that future resilience in food production systems will require more explicit links between consumers and the work of supporting ecosystems, locally and in other regions of the world, and that food security planning will require active management of the capacity of all involved ecosystems to sustain food production.
|
|
| 14. |
|
|
| 15. |
- Dube, Faruk, et al.
(författare)
-
Benzylpenicillin-producing Trichophyton erinacei and methicillin resistant Staphylococcus aureus carrying the mecC gene on European hedgehogs : a pilot-study
- 2021
-
Ingår i: BMC Microbiology. - : BioMed Central (BMC). - 1471-2180. ; 21
-
Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: A high carriage rate of methicillin-resistant Staphylococcus aureus with the mecC gene (mecC-MRSA) has been described among Wild European hedgehogs (Europeaus erineaus). Due to this frequent occurrence, it has been suggested that hedgehogs could be a natural reservoir for mecC-MRSA. However, the reason why hedgehogs carry mecC-MRSA remains unknown, but it has been hypothesized that mecC-MRSA could have evolved on the skin of hedgehogs due to the co-occurrence with antibiotic producing dermatophytes. The aim of this pilot-study was therefore to investigate if hedgehogs in Sweden carry Trichophyton spp. and to provide evidence that these dermatophytes are able to produce penicillin or similar substances. In addition, the study aimed to identify if dermatophytes co-occurred with mecC-MRSA.METHODS: Samples were collected from hedgehogs (Europeaus erineaus) that were euthanized or died of natural causes. All samples were screened for dermatophytes and mecC-MRSA using selective cultivation methods. Suspected isolates were characterized using PCR-based methods, genome sequencing and bioinformatic analyses. Identification of penicillin was performed by ultra-high-performance liquid chromatography-tandem mass spectrometry.RESULTS: In total 23 hedgehogs were investigated, and it was shown that two carried Trichophyton erinacei producing benzyl-penicillin, and that these hedgehogs also carried mecC-MRSA. The study also showed that 60% of the hedgehogs carried mecC-MRSA.CONCLUSION: The pilot-study demonstrated that Trichophyton erinacei, isolated from Swedish hedgehogs, can produce benzylpenicillin and that these benzylpenicillin-producing T. erinacei co-occurred with mecC-MRSA. The study also reconfirmed the high occurrence of mecC-MRSA among hedgehogs.
|
|
| 16. |
- Einarsson, Elin, et al.
(författare)
-
Coordinated Changes in Gene Expression Throughout Encystation of Giardia intestinalis
- 2016
-
Ingår i: PLoS Neglected Tropical Diseases. - : Public Library of Science (PLoS). - 1935-2727 .- 1935-2735. ; 11:3
-
Tidskriftsartikel (refereegranskat)abstract
- Differentiation into infectious cysts through the process of encystation is crucial for transmission and survival of the intestinal protozoan parasite Giardia intestinalis. Hitherto the majority of studies have focused on the early events, leaving late encystation poorly defined. In order to further study encystation, focusing on the later events, we developed a new encystation protocol that generates a higher yield of mature cysts compared to standard methods. Transcriptome changes during the entire differentiation from trophozoites to cysts were thereafter studied using RNA sequencing (RNA-seq). A high level of periodicity was observed for up-and down-regulated genes, both at the level of the entire transcriptome and putative regulators. This suggests the trajectory of differentiation to be coordinated through developmentally linked gene regulatory activities. Our study identifies a core of 13 genes that are consistently up-regulated during initial encystation. Of these, two constitute previously uncharacterized proteins that we were able to localize to a new type of encystation-specific vesicles. Interestingly, the largest transcriptional changes were seen in the late phase of encystation with the majority of the highly up-regulated genes encoding hypothetical proteins. Several of these were epitope-tagged and localized to further characterize these previously unknown genetic components of encystation and possibly excystation. Finally, we also detected a switch of variant specific surface proteins (VSPs) in the late phase of encystation. This occurred at the same time as nuclear division and DNA replication, suggesting a potential link between the processes.
|
|
| 17. |
- Einarsson, Elin, et al.
(författare)
-
UV irradiation responses in Giardia intestinalis
- 2015
-
Ingår i: Experimental parasitology. - : Elsevier BV. - 0014-4894 .- 1090-2449. ; 154, s. 25-32
-
Tidskriftsartikel (refereegranskat)abstract
- The response to ultraviolet light (UV) radiation, a natural stressor to the intestinal protozoan parasite Giardia intestinalis, was studied to deepen the understanding of how the surrounding environment affects the parasite during transmission. UV radiation at 10 mJ/cm(2) kills Giardia cysts effectively whereas trophozoites and encysting parasites can recover from UV treatment at 100 mJ/cm(2) and 50 mJ/cm(2) respectively. Staining for phosphorylated histone H2A showed that UV treatment induces double-stranded DNA breaks and flow cytometry analyses revealed that UV treatment of trophozoites induces DNA replication arrest. Active DNA replication coupled to DNA repair could be an explanation to why UV light does not kill trophozoites and encysting cells as efficiently as the non-replicating cysts. We also examined UV-induced gene expression responses in both trophozoites and cysts using RNA sequencing (RNA seq). UV radiation induces small overall changes in gene expression in Giardia but cysts show a stronger response than trophozoites. Heat shock proteins, kinesins and Nek kinases are up-regulated, whereas alpha-giardins and histones are down-regulated in UV treated trophozoites. Expression of variable surface proteins (VSPs) is changed in both trophozoites and cysts. Our data show that Giardia cysts have limited ability to repair UV-induced damage and this may have implications for drinking- and waste-water treatment when setting criteria for the use of UV disinfection to ensure safe water.
|
|
| 18. |
- Haxton, Eva, et al.
(författare)
-
One Health education meets science
- 2015
-
Ingår i: Infection Ecology & Epidemiology. - Uppsala : Informa UK Limited. - 2000-8686. ; 5
-
Tidskriftsartikel (övrigt vetenskapligt/konstnärligt)
|
|
| 19. |
- Hofstetrova, Klara, et al.
(författare)
-
Giardia intestinalis : Aphidicolin influence on the trophozoite cell cycle
- 2010
-
Ingår i: Experimental parasitology. - : Elsevier BV. - 0014-4894 .- 1090-2449. ; 124:2, s. 159-166
-
Tidskriftsartikel (refereegranskat)abstract
- This study is a thorough examination of the effects of the DNA polymerase inhibitor aphidicolin on the nuclear cycle and cell cycle progression characteristics, as well as their reversibility, in Giardia intestinalis. Giardia trophozoites are arrested in the G1/S-junction after aphidicolin treatment according to their DNA content. However, cell growth continues and trophozoites arrested with aphidicolin resemble cells in the G2 phase and trophozoites in ageing cultures. Extensive treatment with aphidicolin causes side effects and we detected positive signals for phosphorylated histone H2A, which, in mammalian cells. is involved in a signalling pathway triggered as a reaction to double stranded DNA breaks. These results suggest that aphidicolin causes dissociation of the nuclear and cytoplasmic cycles, a phenomenon that has also been described for other inhibitors in mammalian cell lines. Thus, if aphidicolin is used for synchronization of Giardia trophozoites, this fact must be accounted for, and treatment with aphidicolin must be minimal. (C) 2009 Elsevier Inc. All rights reserved.
|
|
| 20. |
- Klotz, Christian, et al.
(författare)
-
Extensive testing of a multi-locus sequence typing scheme for Giardia duodenalis assemblage A confirms its good discriminatory power
- 2022
-
Ingår i: Parasites & Vectors. - : BioMed Central (BMC). - 1756-3305. ; 15
-
Tidskriftsartikel (refereegranskat)abstract
- Background: The flagellated parasite Giardia duodenalis is a major and global cause of diarrhoeal disease. Eight genetically very distinct groups, known as assemblages A to H, have been recognized in the G. duodenalis species complex, two of which (assemblages A and B) infect humans and other mammalian hosts. Informative typing schemes are essential to understand transmission pathways, characterize outbreaks and trace zoonotic transmission. In this study, we evaluated a published multi-locus sequence typing (MLST) scheme for G. duodenalis assemblage A, which is based on six polymorphic markers.Methods: We genotyped 60 human-derived and 11 animal-derived G. duodenalis isolates collected in Europe and on other continents based on the published protocol. After retrieving previously published genotyping data and excluding isolates whose sequences showed allelic sequence heterozygosity, we analysed a dataset comprising 146 isolates.Results: We identified novel variants at five of the six markers and identified 78 distinct MLST types in the overall dataset. Phylogenetic interpretation of typing data confirmed that sub-assemblage AII only comprises human-derived isolates, whereas sub-assemblage AI comprises all animal-derived isolates and a few human-derived isolates, suggesting limited zoonotic transmission. Within sub-assemblage AII, isolates from two outbreaks, which occurred in Sweden and Italy, respectively, had unique and distinct MLST types. Population genetic analysis showed a lack of clustering by geographical origin of the isolates.Conclusion: The MLST scheme evaluated provides sufficient discriminatory power for epidemiological studies of G. duodenalis assemblage A.
|
|
| 21. |
- Lilja, Tobias, et al.
(författare)
-
Species identification of Swedish mosquitoes through DNA metabarcoding
- 2017
-
Ingår i: Journal of the European Mosquito Control Association. ; 35, s. 1-9
-
Tidskriftsartikel (refereegranskat)abstract
- DNA-barcoding utilises a fragment of the mitochondrial cytochrome oxidase subunit 1 (COI) gene to identify most animal species. Using next generation sequencing (NGS), this method can be further developed into metabarcoding processes that allow the simultaneous identification of several species from a mixed sample. We created a database of COI sequences of 27 mosquito species collected in Sweden, and combined our data with 27 additional sequences from GenBank to cover the taxa recently documented in Sweden and to include possible invasive taxa. Comparisons show that COI metabarcoding reliably identifies 41 of 54 species and the remainder to species group. Using three independent primer pairs along the COI gene, we further developed this barcoding approach to simultaneously identify Swedish mosquitoes in communities using NGS and quantify relative abundance of each mosquito species in the sample, using bioinformatics methods. We tested the accuracy of the metabarcoding method using communities assembled from morphologically identified mosquitoes, revealing 80% positive identification rate and the estimates of population structure which reflects the input sample. We conclude that metabarcoding is useful as a high throughput identification technique and for the quantification of species.
|
|
| 22. |
- Ngubane, Zesizwe, et al.
(författare)
-
Water quality modelling and quantitative microbial risk assessment for uMsunduzi River in South Africa
- 2022
-
Ingår i: Journal of Water and Health. - : IWA Publishing. - 1996-7829 .- 1477-8920. ; 20:4, s. 641-656
-
Tidskriftsartikel (refereegranskat)abstract
- South African rivers generally receive waste from inadequate wastewater infrastructure, mines, and farming activities, among others. The uMsunduzi River in KwaZulu-Natal, South Africa, is among these recipients with recorded poor to very poor water quality. To identify parts of the uMsunduzi River that are polluted by Cryptosporidium and Escherichia coli (E. coli), this study mapped out pollutants emanating from point and non-point sources using the Soil and Water Assessment Tool (SWAT). Streamflow calibration in the upper and lower reaches of the catchment showed good performance with R-2 of 0.64 and 0.58, respectively. SWAT water quality output data were combined with a Quantitative Microbial Risk Assessment (QMRA) to understand the microbial health implications for people using river water for drinking, recreational swimming, and non-competitive canoeing. QMRA results for Cryptosporidium and pathogenic E. coli showed that the probability of infection for most users exceeds the acceptable level for drinking and recreation as outlined in the South African water quality guidelines, and by the World Health Organisation (WHO). The results of this study can be used as a baseline to assess the economic and health implications of different management plans, resulting in better-informed, cost-effective, and impactful decision-making.
|
|
| 23. |
- Reiner, David S., et al.
(författare)
-
Synchronisation of Giardia lamblia : identification of cell cycle stage-specific genes and a differentiation restriction point
- 2008
-
Ingår i: International Journal of Parasitology. - : Elsevier BV. - 0020-7519 .- 1879-0135. ; 38:8-9, s. 935-944
-
Tidskriftsartikel (refereegranskat)abstract
- The intestinal parasite Giardia lamblia undergoes cell differentiations that entail entry into and departure from the replicative cell cycle. The pathophysiology of giardiasis depends directly upon the ability of the trophozoite form to replicate in the host upper small intestine. Thus, cell proliferation is tightly linked to disease. However, studies of cell cycle regulation in Giardia have been hampered by the inability to synchronise cultures. Here we report that Giardia isolates of the major human genotypes A and B can be synchronised using aphidicolin, a mycotoxin that reversibly inhibits replicative DNA polymerases in eukaryotic cells. Aphidicolin arrests Giardia trophozoites in the early DNA synthesis (S) phase of the cell cycle. We identified a set of cell cycle orthologues in the Giardia genome using bioinformatic analyses and showed that synchronised parasites express these genes in a cell cycle stage-specific manner. The synchronisation method also showed that during encystation, exit from the ordinary cell cycle occurs preferentially in GZ and defines a restriction point for differentiation. Synchronisation opens up possibilities for further molecular and cell biological studies of chromosome replication, mitosis and segregation of the complex cytoskeleton in Giardia.
|
|
| 24. |
- Rojas-Lopez, Laura, et al.
(författare)
-
Development of a gp60-subtyping method for Cryptosporidium felis
- 2020
-
Ingår i: Parasites & Vectors. - : BMC. - 1756-3305. ; 13
-
Tidskriftsartikel (refereegranskat)abstract
- Background: Feline cryptosporidiosis is an increasing problem, especially in catteries. In humans, close contact with cats could be a potential source of infection although the risk of contracting cryptosporidiosis caused by Cryptosporidium felis is considered to be relatively low. Sequencing of the 60-kDa glycoprotein gene is a commonly used tool for investigation of the genetic diversity and transmission dynamics of Cryptosporidium species. However, until now the sequence of gp60 from C. felis has not been available and genotyping has been limited to less discriminatory markers, such as 18S rRNA, COWP and HSP70.Methods: We have identified the gp60 orthologue within the genome sequence of C. felis, and used the sequence to design a nested PCR for subtyping purposes. A total of 128 clinical isolates of both feline and human origin, were used to evaluate the marker.Results: Sequence analysis revealed large variations between the different samples. The C. felis gp60 lack the characteristic serine-tract found in many other cryptosporidian orthologues, instead it has an insertion of variable length (361-742 nt). Also, two cases of suspected zoonotic transmission of C. felis between cats and humans were successfully confirmed.Conclusions: We have identified the gp60 gene in C. felis and show how this highly variable marker can be used in epidemiological investigations.
|
|
| 25. |
- Roxström-Lindquist, Katarina, et al.
(författare)
-
Large genomic differences between the morphologically indistinguishable diplomonads Spironucleus barkhanus and Spironucleus salmonicida.
- 2010
-
Ingår i: BMC Genomics. - 1471-2164. ; 11, s. 258-
-
Tidskriftsartikel (refereegranskat)abstract
- Background Microbial eukaryotes show large variations in genome structure and content between lineages, indicating extensive flexibility over evolutionary timescales. Here we address the tempo and mode of such changes within diplomonads, flagellated protists with two nuclei found in oxygen-poor environments. Approximately 5,000 expressed sequence tag (EST) sequences were generated from the fish commensal Spironucleus barkhanus and compared to sequences from the morphologically indistinguishable fish parasite Spironucleus salmonicida, and other diplomonads. The ESTs were complemented with sequence variation studies in selected genes and genome size determinations. Results Many genes detected in S. barkhanus and S. salmonicida are absent in the human parasite Giardia intestinalis, the most intensively studied diplomonad. For example, these fish diplomonads show an extended metabolic repertoire and are able to incorporate selenocysteine into proteins. The codon usage is altered in S. barkhanus compared to S. salmonicida. Sequence variations were found between individual S. barkhanus ESTs for many, but not all, protein coding genes. Conversely, no allelic variation was found in a previous genome survey of S. salmonicida. This difference was confirmed by sequencing of genomic DNA. Up to five alleles were identified for the cloned S. barkhanus genes, and at least nineteen highly expressed S. barkhanus genes are represented by more than four alleles in the EST dataset. This could be explained by the presence of a non-clonal S. barkhanus population in the culture, by a ploidy above four, or by duplications of parts of the genome. Indeed, genome size estimations using flow cytometry indicated similar haploid genome sizes in S. salmonicida and G. intestinalis (similar to 12 Mb), whereas the S. barkhanus genome is larger (similar to 18 Mb). Conclusions This study indicates extensive divergent genome evolution within diplomonads. Genomic traits such as codon usage, frequency of allelic sequence variation, and genome size have changed considerably between S. barkhanus and S. salmonicida. These observations suggest that large genomic differences may accumulate in morphologically indistinguishable eukaryotic microbes.
|
|
| 26. |
- Sandholt, Arnar K. S., et al.
(författare)
-
Dual RNA-seq transcriptome analysis of caecal tissue during primary Eimeria tenella infection in chickens
- 2021
-
Ingår i: BMC Genomics. - : BioMed Central (BMC). - 1471-2164. ; 22
-
Tidskriftsartikel (refereegranskat)abstract
- Background: Coccidiosis is an infectious disease with large negative impact on the poultry industry worldwide. It is an enteric infection caused by unicellular Apicomplexan parasites of the genus Eimeria. The present study aimed to gain more knowledge about interactions between parasites and the host immune system during the early asexual replication phase of E. tenella in chicken caeca. For this purpose, chickens were experimentally infected with E. tenella oocysts, sacrificed on days 1-4 and 10 after infection and mRNA from caecal tissues was extracted and sequenced.Results: Dual RNA-seq analysis revealed time-dependent changes in both host and parasite gene expression during the course of the infection. Chicken immune activation was detected from day 3 and onwards with the highest number of differentially expressed immune genes recorded on day 10. Among early (days 3-4) responses up-regulation of genes for matrix metalloproteinases, several chemokines, interferon (IFN)-gamma along with IFN-stimulated genes GBP, IRF1 and RSAD2 were noted. Increased expression of genes with immune suppressive/regulatory effects, e.g. IL10, SOCS1, SOCS3, was also observed among early responses. For E. tenella a general up-regulation of genes involved in protein expression and energy metabolism as well as a general down-regulation genes for DNA and RNA processing were observed during the infection. Specific E. tenella genes with altered expression during the experiment include those for proteins in rhoptry and microneme organelles.Conclusions: The present study provides novel information on both the transcriptional activity of E. tenella during schizogony in ceacal tissue and of the local host responses to parasite invasion during this phase of infection. Results indicate a role for IFN-gamma and IFN-stimulated genes in the innate defence against Eimeria replication.
|
|
| 27. |
- Sandholt, Arnar K. S., et al.
(författare)
-
Dual RNA-Seq transcriptome analysis of chicken macrophage-like cells (HD11) infected in vitro with Eimeria tenella
- 2021
-
Ingår i: Parasitology. - : Cambridge University Press. - 0031-1820 .- 1469-8161. ; 148:6, s. 712-725
-
Tidskriftsartikel (refereegranskat)abstract
- The study aimed to monitor parasite and host gene expression during the early stages of Eimeria tenella infection of chicken cells using dual RNA-Seq analysis. For this, we used chicken macrophage-like cell line HD11 cultures infected in vitro with purified E. tenella sporozoites. Cultures were harvested between 2 and 72 h post-infection and mRNA was extracted and sequenced. Dual RNA-Seq analysis showed clear patterns of altered expression for both parasite and host genes during infection. For example, genes in the chicken immune system showed upregulation early (2-4 h), a strong downregulation of genes across the immune system at 24 h and a repetition of early patterns at 72 h, indicating that invasion by a second generation of parasites was occurring. The observed downregulation may be due to immune self-regulation or to immune evasive mechanisms exerted by E. tenella. Results also suggested pathogen recognition receptors involved in E. tenella innate recognition, MRC2, TLR15 and NLRC5 and showed distinct chemokine and cytokine induction patterns. Moreover, the expression of several functional categories of Eimeria genes, such as rhoptry kinase genes and microneme genes, were also examined, showing distinctive differences which were expressed in sporozoites and merozoites.
|
|
| 28. |
- Sanno, Axel, et al.
(författare)
-
Sarcoptic mange in the wild boar, Sus scrofa, in Sweden
- 2021
-
Ingår i: CURRENT RESEARCH IN PARASITOLOGY & VECTOR-BORNE DISEASES. - : Elsevier. - 2667-114X. ; 1
-
Tidskriftsartikel (refereegranskat)abstract
- Sarcoptic mange caused by Sarcoptes scabiei has been present in the Swedish red fox (Vulpes vulpes) population since the 1970s. The disease has been described in other Swedish wildlife species, but not in the wild boar, Sus scrofa, until 2009. Single cases of sarcoptic mange have been diagnosed the last years in the expanding population of wild boar. This study aims to describe the histopathological lesions found on mangy wild boar and compare, by molecular methods, mites from wild boar cases with mites from mangy red foxes, raccoon dogs, and domestic pigs. Mangy wild boar with focal alopecia and clinical signs of pruritis were reported or submitted from various areas in southern Sweden to the National Veterinary Institute, Uppsala. The examined skin samples of wild boar infected with S. scabiei showed limited gross skin lesions, except for cases with severe exudative dermatitis. Histopathology of the affected wild boar skin samples showed an eosinophilic dermatitis with a variable hyperkeratosis and often low number of mites present. To study the relationship of S. scabiei mites isolated from different host species, a population genetics investigation was performed based on microsatellite markers. In total, 225 individual mites from eight individuals of four different host species; red fox (48 mites), wild boar (80 mites), domestic pig (48 mites) and raccoon dog (43 mites), were included in the study. In the phylogenetic analysis, all mites isolated from wild boar clustered together even though they originate from different geographical regions in Sweden. Mites from each individual host showed high similarity. The results indicate that wild boar mites differ from mites both from the red fox, raccoon dog, and domestic pig.
|
|
| 29. |
- Sikora, Per, et al.
(författare)
-
Genomic Variation in IbA10G2 and Other Patient-Derived Cryptosporidium hominis Subtypes
- 2017
-
Ingår i: Journal of Clinical Microbiology. - : AMER SOC MICROBIOLOGY. - 0095-1137 .- 1098-660X. ; 55:3, s. 844-858
-
Tidskriftsartikel (refereegranskat)abstract
- In order to improve genotyping and epidemiological analysis of Cryptosporidium spp., genomic data need to be generated directly from a broad range of clinical specimens. Utilizing a robust method that we developed for the purification and generation of amplified target DNA, we present its application for the successful isolation and whole-genome sequencing of 14 different Cryptosporidium hominis patient specimens. Six isolates of subtype IbA10G2 were analyzed together with a single representative each of 8 other subtypes: IaA20R3, IaA23R3, IbA9G3, IbA13G3, IdA14, IeA11G3T3, IfA12G1, and IkA18G1. Parasite burden was measured over a range of more than 2 orders of magnitude for all samples, while the genomes were sequenced to mean depths of between 17X and 490X coverage. Sequence homologybased functional annotation identified several genes of interest, including the gene encoding Cryptosporidium oocyst wall protein 9 (COWP9), which presented a predicted loss-of-function mutation in all the sequence subtypes, except for that seen with IbA10G2, which has a sequence identical to the Cryptosporidium parvum reference Iowa II sequence. Furthermore, phylogenetic analysis showed that all the IbA10G2 genomes form a monophyletic clade in the C. hominis tree as expected and yet display some heterogeneity within the IbA10G2 subtype. The current report validates the aforementioned method for isolating and sequencing Cryptosporidium directly from clinical stool samples. In addition, the analysis demonstrates the potential in mining data generated from sequencing multiple whole genomes of Cryptosporidium from human fecal samples, while alluding to the potential for a higher degree of genotyping within Cryptosporidium epidemiology.
|
|
| 30. |
- Sumaila, U. Rashid, et al.
(författare)
-
WTO must ban harmful fisheries subsidies
- 2021
-
Ingår i: Science. - : American Association for the Advancement of Science (AAAS). - 0036-8075 .- 1095-9203. ; 374:6567, s. 544-544
-
Tidskriftsartikel (övrigt vetenskapligt/konstnärligt)
|
|
| 31. |
- Søgaard Jørgensen, Peter, et al.
(författare)
-
Coevolutionary Governance of Antibiotic and Pesticide Resistance
- 2020
-
Ingår i: Trends in Ecology & Evolution. - : Elsevier BV. - 0169-5347 .- 1872-8383. ; 35:6, s. 484-494
-
Forskningsöversikt (refereegranskat)abstract
- Development of new biocides has dominated human responses to evolution of antibiotic and pesticide resistance. Increasing and uniform biocide use, the spread of resistance genes, and the lack of new classes of compounds indicate the importance of navigating toward more sustainable coevolutionary dynamics between human culture and species that evolve resistance. To inform this challenge, we introduce the concept of coevolutionary governance and propose three priorities for its implementation: (i) new norms and mental models for lowering use, (ii) diversifying practices to reduce directional selection, and (iii) investment in collective action institutions to govern connectivity. We highlight the availability of solutions that facilitate broader sustainable development, which for antibiotic resistance include improved sanitation and hygiene, strong health systems, and decreased meat consumption.
|
|
| 32. |
- Troell, Karin, et al.
(författare)
-
Cryptosporidium as a testbed for single cell genome characterization of unicellular eukaryotes
- 2016
-
Ingår i: BMC Genomics. - : Springer Science and Business Media LLC. - 1471-2164. ; 17
-
Tidskriftsartikel (refereegranskat)abstract
- Background: Infectious disease involving multiple genetically distinct populations of pathogens is frequently concurrent, but difficult to detect or describe with current routine methodology. Cryptosporidium sp. is a widespread gastrointestinal protozoan of global significance in both animals and humans. It cannot be easily maintained in culture and infections of multiple strains have been reported. To explore the potential use of single cell genomics methodology for revealing genome-level variation in clinical samples from Cryptosporidium-infected hosts, we sorted individual oocysts for subsequent genome amplification and full-genome sequencing. Results: Cells were identified with fluorescent antibodies with an 80 % success rate for the entire single cell genomics workflow, demonstrating that the methodology can be applied directly to purified fecal samples. Ten amplified genomes from sorted single cells were selected for genome sequencing and compared both to the original population and a reference genome in order to evaluate the accuracy and performance of the method. Single cell genome coverage was on average 81 % even with a moderate sequencing effort and by combining the 10 single cell genomes, the full genome was accounted for. By a comparison to the original sample, biological variation could be distinguished and separated from noise introduced in the amplification. Conclusions: As a proof of principle, we have demonstrated the power of applying single cell genomics to dissect infectious disease caused by closely related parasite species or subtypes. The workflow can easily be expanded and adapted to target other protozoans, and potential applications include mapping genome-encoded traits, virulence, pathogenicity, host specificity and resistance at the level of cells as truly meaningful biological units.
|
|
| 33. |
- Troell, Karin, et al.
(författare)
-
Cryptosporidium parvum infections in a cohort of veterinary students in Sweden
- 2015
-
Ingår i: Epidemiology and Infection. - 0950-2688 .- 1469-4409. ; 143, s. 2748-2756
-
Tidskriftsartikel (refereegranskat)abstract
- In March 2013, a veterinary student tested positive for Cryptosporidium; four classmates reported similar gastrointestinal symptoms. We aimed to identify source(s) and risk factors for Cryptosporidium infection in university persons symptomatic between 21 January and 14 April 2013. Sixty-four (79%) students from a cohort of 81 fourth-year veterinary students completed questionnaires, identifying 13 cases; four were Cryptosporidium parvum GP60 subtype IIaA16G1R1b, two were IIdA24G1, seven did not submit stool samples. Thirteen cases attended the university's field clinic before symptom onset (13/37 attendees, 35%); 11 visited at least one of four farms where students recalled seeing calves with diarrhoea. C. parvum subtype IIaA16G1R1b was identified in calves at one of the farms. Entering pens of calves with diarrhoea [relative risk (RR) 7.6, 95% confidence interval (CI) 1.7-33.5] and eating in clinic cars (RR 9.1, 95% CI 1.3-65.8) were associated with being a case. Washing hands at least twice per farm visit (0 cases, P = 0.03) was protective. This outbreak investigation was notable for rapid and effective collaboration between public health, veterinary and environmental sectors, leading to swift identification of a microbiological and epidemiological link between cases, infected calves and their farms. We recommend frequent hand-washing using proper technique and dissuasion from eating in clinic cars to minimize possible exposure to contaminated surfaces.
|
|
| 34. |
- Yanta, Christine A., et al.
(författare)
-
CryptoGenotyper : A new bioinformatics tool for rapid Cryptosporidium identification
- 2021
-
Ingår i: FOOD AND WATERBORNE PARASITOLOGY. - : Elsevier. - 2405-6766. ; 23
-
Tidskriftsartikel (refereegranskat)abstract
- Cryptosporidium is a protozoan parasite that is transmitted to both humans and animals through zoonotic or anthroponotic means. When a host is infected with this parasite, it causes a gastrointestinal disease known as cryptosporidiosis. To understand the transmission dynamics of Cryptosporidium, the small subunit (SSU or 18S) rRNA and gp60 genes are commonly studied through PCR analysis and conventional Sanger sequencing. However, analyzing sequence chromatograms manually is both time consuming and prone to human error, especially in the presence of poorly resolved, heterozygous peaks and the absence of a validated database. For this study, we developed a Cryptosporidium genotyping tool, called CryptoGenotyper, which has the capability to read raw Sanger sequencing data for the two common Cryptosporidium gene targets (SSU rRNA and gp60) and classify the sequence data into standard nomenclature. The CryptoGenotyper has the capacity to perform quality control and properly classify sequences using a high quality, manually curated reference database, saving users' time and removing bias during data analysis. The incorporated heterozygous base calling algorithms for the SSU rRNA gene target resolves double peaks, therefore recovering data previously classified as inconclusive. The CryptoGenotyper successfully genotyped 99.3% (428/431) and 95.1% (154/162) of SSU rRNA chromatograms containing single and mixed sequences, respectively, and correctly subtyped 95.6% (947/991) of gp60 chromatograms without manual intervention. This new, user-friendly tool can provide both fast and reproducible analyses of Sanger sequencing data for the two most common Cryptosporidium gene targets. Crown Copyright (C) 2021 Published by Elsevier Inc. on behalf of International Association of Food and Waterborne Parasitology.
|
|
