First detection of Ichthyophonus sp. in invasive wild pink salmon (Oncorhynchus gorbuscha) from the North Atlantic Ocean.

Pacific pink salmon (Oncorhynchus gorbuscha) were deliberately introduced to rivers surrounding the White Sea and has spread to Norway and several other countries surrounding the North Atlantic Ocean. In August 2021, a female pink salmon displaying pale gills and abnormal behaviour was captured in River Lakselva in Northern Norway and later submitted to the Norwegian Veterinary Institute (NVI) for post-mortem examination. Histological examination of organ samples revealed structures indicative of systemic ichthyophoniasis, caused by Ichthyophonus sp. The parasites appeared to be especially abundant in the heart and skeletal musculature, and local tissue responses were assessed to be absent or very mild. Sequences of the ribosomal 18S rRNA and the mitochondrial cytochrome oxidase 1 (CO1) genes confirmed the diagnosis and identified the pathogen as Ichthyophonus sp. The CO1 sequence further established that the isolate from pink salmon was most similar to sequences of Ichthyophonus sp. from Atlantic salmon, Salmo salar, from the Atlantic Ocean on the east coast of the US and from Atlantic herring, Clupea harengus, from Iceland. We here report the first detection of Ichthyophonus sp. in pink salmon in the North Atlantic Ocean.

Non-lethal detection of Eubothrium crassum (Cestoda) in farmed Atlantic salmon, Salmo salar, using anal swabs and real-time PCR.

Detection of intestinal parasites in fish typically requires autopsy, resulting in the sacrifice of the fish. Here, we describe a non-lethal method for detecting the tapeworm Eubothrium crassum in fish using anal swabs and real-time PCR detection. Two assays were developed to detect cytochrome oxidase I (COI) mitochondrial DNA and 18S ribosomal DNA sequences of E. crassum, respectively. The assays were tested on swab samples from confirmed pathogen free Atlantic salmon (Salmo salar L.) and on samples from farmed Atlantic salmon, where the presence and intensity of parasites had been established through autopsy. The COI assay was shown to be specific to E. crassum, while the 18S assay also amplified the closely related E. salvelini, a species infecting Arctic charr (Salvelinus alpinus L.) in freshwater. The COI assay detected E. crassum in all field samples regardless of parasite load while the 18S assay failed to detect the parasite in two samples. The results thus demonstrates that this non-lethal approach can effectively detect E. crassum and can be a valuable tool in assessing the prevalence of infection in farmed salmon, aiding in treatment decisions and evaluating treatment effectiveness.

Proposal of Troglocephalinae n. subfam. (Monogenea: Monocotylidae) to accommodate existing and two new monocotylids from the gills of rhinopristiform shovelnose rays.

Troglocephalinae n. subfam. is proposed for Spinuris Doran, 1953, Neoheterocotyle Hargis, 1955, Anoplocotyloides Young, 1967, Troglocephalus rhinobatidis Young, 1967 (previously incertae sedis), Nonacotyle pristis Ogawa, 1991, Mehracotyle insolita Neifar, Euzet & Ben Hassine, 2002, Scuticotyle cairae n. gen. et sp., and Brancheocotyle imbricata n. gen. et sp. All members of the proposed new subfamily are gill parasites of shovelnose rays of the order Rhinopristiformes. The subfamilies Heterocotylinae Chisholm, Wheeler & Beverley-Burton, 1995, and Dasybatotreminae Bychowsky, 1957, are amended to exclude Spinuris, Nonacotyle, Neoheterocotyle, and Anoplocotyloides and Mehracotyle, respectively. Heterocotylinae includes gill parasites of members of the orders Myliobatiformes and Torpediniformes. Dasybatotreminae includes parasites of the gills and pharyngeal cavity of members of the orders Myliobatiformes and Rajiformes. A revised phylogeny of the Monocotylidae Taschenberg, 1879 is presented and discussed, based on 28S rDNA sequences, including new sequences for Myliocotyle pteromylaei Neifer, Euzet & Ben Hassine, 1999, Heterocotyle tokoloshei Vaughan & Chisholm, 2010, Neoheterocotyle robii Vaughan & Chisholm, 2010, and the two newly proposed species and genera. Additional locality records are also provided for Monocotylidae from off South Africa.

Occurrence and seasonality of Gyrodactylus salaris and G. salmonis (Monogenea) on Arctic char (Salvelinus alpinus (L.)) in the Fustvatnet lake, Northern Norway.

Gyrodactylus spp. (Monogenea) were found on 16.9% (233 out of 1376) Arctic char, Salvelinus alpinus (L.), sampled from September 2010 to October 2011 in the Fustvatnet lake, Northern Norway. Two species were identified: G. salaris Malmberg, 1957, and G. salmonis Yin & Sproston, 1948. Gyrodactylus salaris was only found on Arctic char larger than 28 cm and only in samples obtained in the autumn (September and October). Gyrodactylus salmonis was found on Arctic char of all sizes (11-47 cm) and throughout the year, with a small peak in abundance in the late autumn (November). Gyrodactylus salaris was found to prefer the tail and dorsal fin. Based on the results, we recommend that surveys of Arctic char for the presence of G. salaris are based on the examination of the fins of large fish sampled during the spawning season (autumn).

New monogeneans from the bathydemersal southern African endemic catshark, Holohalaelurus regani (Gilchrist, 1922).

The catshark genus Holohalaelurus Fowler is currently represented by five species distributed off the southern and eastern parts of the African coast. Very few parasitological records exist for any of these five species, representing a significant knowledge gap. We report the first monogenean species from the bathydemersal species Holohalaelurus regani (Gilchrist): a new species of Microbothriidae Price, 1936, Leptomicrobothrium holohalaelure n. sp. from the dorsal skin surface, and a new species of Hexabothriidae Price, 1942 representing a new genus, Scyliorhinocotyle narvaezae n. gen., n. sp., from the gill lamellae. Both monogenean records represent the first for any member of the catshark genus Holohalaelurus. Previously we focused on the identification of monogenean taxa of emerging veterinary importance for public aquaria. We now begin a focused effort to document a generally unexplored monogenean biodiversity from diverse marine habitats off South Africa.

Genetic diversity among geographically distant isolates of Neoparamoeba perurans.

The present study explored the use of 2 common genetic markers, the mitochondrial cytochrome oxidase I (COI) and the ribosomal internal transcribed spacer (ITS) to infer the relationship between geographically distant isolates of the protozoan gill parasite Neoparamoeba perurans, the agent responsible for amoebic gill disease in farmed Atlantic salmon worldwide. Present data confirmed that the ITS marker is suitable for Neoparamoeba species discrimination; however, it is not recommended as a population marker due to the presence of multiple copies of ITS within both N. perurans clonal and polycultures. On the other hand, in the partial COI gene analysed here, a low variability was observed, with 8 haplotypes recovered from N. perurans samples collected from Europe (Ireland, Norway, Scotland) and Tasmania (Australia). In Europe, the COI haplotypes which have more recently been detected in aquaculture are different to the haplotypes associated with the original gill disease emergence in Ireland in 1997 and Norway in 2006. The presence of unique COI haplotypes in different continents suggests the presence of multiple distinct reservoirs of the pathogen in both Europe and Tasmania. Isolates from additional geographical locations are required to fully understand the origins and routes for the spread of N. perurans worldwide.

Systematic screening for atrial fibrillation in a 65-year-old population with risk factors for stroke: data from the Akershus Cardiac Examination 1950 study.

Aims: To investigate the yield of screening for atrial fibrillation (AF) in a cohort of 65-year-old individuals from the general population with additional risk factors for stroke. Methods and results: We invited participants with additional risk factors for stroke (CHA2DS2-VASc score ≥2 for men or ≥ 3 for women) without previously known AF from a population-based study in Norway to participate in a 2-week screening for AF. Screening was performed by one-lead 'thumb electrocardiography (ECG)' recordings of 30 s twice daily or when the participants experienced symptoms. In total, 1742 (47.0%) participants of the Akershus Cardiac Examination (ACE) 1950 study had at least one additional risk factor for stroke. Of these, 123 cases reported a history of AF and 101 (5.8%) cases were ECG validated. Eight [0.5%, 95% confidence interval (CI) 0.2-0.9] new AF cases were diagnosed by 12-lead ECG at baseline, and 10 additional participants were diagnosed with AF before screening commenced. We invited all 1601 participants who met the inclusion criteria for screening, of which 1510 (94.3%) participants were included (44% women and 56% men). The screening revealed AF in 13 (0.9%, 95% CI 0.5-1.5) participants. The total prevalence of ECG-validated AF after screening among the 65-year-olds with risk factors for stroke was 7.6% (95% CI 6.4-8.9), in men 10.0% (95% CI 8.2-12.0), and in women 4.3% (95% CI 3.0-6.1) (P < 0.001). Conclusion: In a group of 1510 well-characterized 65-year-olds with risk factors for stroke, 2-week intermittent ECG screening identified undiagnosed AF in 0.9%. The total prevalence of AF was 7.6%.

Integrative Approach for the Reliable Detection and Specific Identification of the Microsporidium Loma morhua in Atlantic Cod (Gadus morhua).

Microsporidia are fungal parasites that infect diverse invertebrate and vertebrate hosts. Finfish aquaculture supports epizootics due to high host density and the high biotic potential of these parasites. Reliable methods for parasite detection and identification are a necessary precursor to empirical assessment of strategies to mitigate the effects of these pathogens during aquaculture. We developed an integrative approach to detect and identify Loma morhua infecting Atlantic cod. We show that the spleen is more reliable than the commonly presumed gills as best organ for parasite detection in spite of substantial morphological plasticity in xenoma complexes. We developed rDNA primers with 100% sensitivity in detecting L. morhua and with utility in distinguishing some congeneric Loma species. ITS sequencing is necessary to distinguish L. morhua from other congeneric microsporidia due to intraspecific nucleotide variation. 64% of L. morhua ITS variants from Atlantic cod have a 9-nucleotide motif that distinguishes it from Loma spp. infecting non-Gadus hosts. The remaining 36% of ITS variants from Atlantic cod are distinguished from currently represented Loma spp., particularly those infecting Gadus hosts, based on a 14-nucleotide motif. This research approach is amenable to developing templates in support of reliable detection and identification of other microsporidian parasites in fishes.

Electrocotyle whittingtoni n. gen., n. sp. (Monogenea: Monocotylidae: Heterocotylinae) from the gills of a captive onefin electric ray, Narke capensis (Narkidae) at Two Oceans Aquarium, Cape Town, South Africa.

Electrocotyle whittingtoni n. gen., n. sp. (Monogenea: Monocotylidae) is described from the gills of a captive female onefin electric ray, Narke capensis, collected for exhibition at Two Oceans Aquarium in Cape Town, South Africa. Electrocotyle n. gen. is most similar to the heterocotyline genera Heterocotyle and Potamotrygonocotyle but could not be accommodated easily in either of these groups. The new genus is characterised by a haptor with one central and eight peripheral loculi, four unsclerotised structures on the dorsal surface of the haptor, a single unsclerotised non-sinous ridge on the ventral surface of the haptoral septa, large hamuli with a long handle and reduced guard, a vagina with sclerotised walls, and tetrahedral eggs. Molecular phylogenetic analyses based on 28S sequences strongly support the separate genus status of Electrocotyle n. gen and thus support our morphological conclusion. The Heterocotylinae is amended to accommodate the new genus, and the new species is fully described and illustrated herein. This is the first record of a monocotylid from the Narkidae. Electrocotyle whittingtoni n. gen. n. sp. is considered potentially pathogenic given its negative impact on the health of its captive host kept in the quarantine facility at Two Oceans Aquarium.

Genetic population structure of Gyrodactylus thymalli (Monogenea) in a large Norwegian river system.

The extent of geographic genetic variation is the result of several processes such as mutation, gene flow, selection and drift. Processes that structure the populations of parasite species are often directly linked to the processes that influence the host. Here, we investigate the genetic population structure of the ectoparasite Gyrodactylus thymalli Zitnan, 1960 (Monogenea) collected from grayling (Thymallus thymallus L.) throughout the river Glomma, the largest watercourse in Norway. Parts of the mitochondrial dehydrogenase subunit 5 (NADH 5) and cytochrome oxidase I (COI) genes from 309 G. thymalli were analysed to study the genetic variation and investigated the geographical distribution of parasite haplotypes. Three main clusters of haplotypes dominated the three distinct geographic parts of the river system; one cluster dominated in the western main stem of the river, one in the eastern and one in the lower part. There was a positive correlation between pairwise genetic distance and hydrographic distance. The results indicate restricted gene flow between sub-populations of G. thymalli, most likely due to barriers that limit upstream migration of infected grayling. More than 80% of the populations had private haplotypes, also indicating long-time isolation of sub-populations. According to a molecular clock calibration, much of the haplotype diversity of G. thymalli in the river Glomma has developed after the last glaciation.

Heart and Brain Interactions--the Akershus Cardiac Examination (ACE) 1950 Study Design.

OBJECTIVES: The aim of the Akershus Cardiac Examination (ACE) 1950 Study is to investigate the development and progression of cardiovascular and cerebrovascular disease (CVD/CeVD) in an extensively characterized age cohort of middle-aged subjects with longitudinal long-term follow-up. DESIGN: The ACE 1950 Study is a prospective, population-based, age-cohort study of all men and women born in 1950 in Akershus County, Norway. The study involves a comprehensive baseline examination, especially for CVD/CeVD, including advanced ultrasound imaging and biobanking ("deep phenotyping"). We expect to obtain an inclusion rate of > 60% from the total study population of 5,827 eligible subjects. Enrollment will be completed during 2015. CONCLUSIONS: The ACE 1950 Study will have potential to generate new and relevant insight into identification of subclinical disease progression. Extensive phenotyping will enable identification of novel disease markers and mechanisms for subclinical disease, which can prove important for future disease prevention.

The precision by the Face Arm Speech Time (FAST) algorithm in stroke capture, sex and age differences: a stroke registry study.

Background: The shift towards milder strokes and studies suggesting that stroke symptoms vary by age and sex may challenge the Face-Arm-Speech Time (FAST) coverage. We aimed to study the proportion of stroke cases admitted with FAST symptoms, sex and age differences in FAST presentation and explore any additional advantage of including new item(s) from the National Institute of Health Stroke Scale (NIHSS) to the FAST algorithm. Methods: This registry-based study included patients admitted with acute stroke to Sahlgrenska University Hospital (November 2014 to June 2019) with NIHSS items at admission. FAST symptoms were extracted from the NIHSS at admission, and sex and age differences were explored using descriptive statistics. Results: Of 5022 patients, 46% were women. Median NIHSS at admission for women was (2 (8-0) and for men 2 (7-0)). In total, 2972 (59%) had at least one FAST symptom, with no sex difference (p=0.22). No sex or age differences were found in FAST coverage when stratifying for stroke severity. 52% suffered mild strokes, whereas 30% had FAST symptoms. The most frequent focal NIHSS items not included in FAST were sensory (29%) and visual field (25%) and adding these or both in modified FAST algorithms led to a slight increase in strokes captured by the algorithms (59%-67%), without providing enhanced prognostic information. Conclusions: 60% had at least one FAST symptom at admission, only 30% in mild strokes, with no sex or age difference. Adding new items from the NIHSS to the FAST algorithm led only to a slight increase in strokes captured.

Can parasites use predators to spread between primary hosts?

Parasites typically have low reproductive fitness on paratenic hosts. Such hosts offer other significant inclusive fitness benefits to parasites, however, such as increased mobility and migration potential. The parasite fauna of the guppy (Poecilia reticulata) is dominated by the directly transmitted ectoparasites Gyrodactylus bullatarudis and Gyrodactylus turnbulli. In the wild, close predatory and competitive interactions occur between the guppy and the killifish Rivulus hartii. Previous observations suggest that these fish can share gyrodactylids, so we tested experimentally whether these parasites can use R. hartii as an alternative host. In aquaria, G. bullatarudis was the only species able to transmit from prey to predator. Both parasite species transferred equally well to prey when the predator was experimentally infected. However, in semi-natural conditions, G. bullatarudis transmitted more successfully to the prey fish. Importantly, G. bullatarudis also survived significantly longer on R. hartii out of water. As R. hartii can migrate overland between isolated guppy populations, G. bullatarudis may have an enhanced ability to disperse and colonize new host populations, consistent with its wider distribution in the wild. To our knowledge, this is the first empirical study demonstrating a predator acting as a paratenic host for the parasites of its prey.

RAJONCHOCOTYLE CERFONTAINE, 1899 (MONOGENEA: HEXABOTHRIIDAE) SPECIES FROM SOUTH AFRICA, WITH DISCUSSION OF THE LITERARY ACCOUNTS OF R. EMARGINATA (OLSSON, 1876).

Rajonchocotyle Cerfontaine, 1899, species are blood-feeding parasites on the gills of rajiforms. Eight species are considered valid, the most recent of which was described just after World War II. Many of the original descriptions of Rajonchocotyle species are limited in diagnostic value, and comparative museum material is scanty. The genus requires revision, and in support thereof, we provide detailed redescriptions for Rajonchocotyle albaCerfontaine, 1899, from the type host Rostroraja alba (Lacepède, 1803) and Rajonchocotyle emarginata (Olsson, 1876), Sproston, 1946, from 2 new host records, Raja straeleni Poll, 1951, and Leucoraja wallacei (Hulley, 1970), from South Africa, a new locality record. The generic diagnosis for Rajonchocotyle is amended to include greater details of the male reproductive system and confirms Paul Cerfontaine's and Nora Sproston's historic observations of the morphology of the male copulatory organ consisting of a separate proximal seminal vesicle and a distal cirrus. The lectotype of Rajonchocotyle kenojeiYamaguti, 1938, is assigned, and a comprehensive Rajonchocotyle-host species list is provided, highlighting species records that require further verification, and the purported global host range of R. emarginata is discussed.

Species of Hexabothriidae (Monogenea) may have extensive distribution ranges reflecting multiple host species: evidence from three new South African records.

Many hexabothriid genera require renewed taxonomic attention to provide additional detailed comparative data from new material to resolve existing species quandaries. Our current study describes and provides additional museum material for three hexabothriid species from three genera from South Africa: Erpocotyle catenulata (Guberlet), which is confirmed as a valid species, Heteronchocotyle gymnurae Neifar, Euzet and Ben Hassine, and Hypanocotyle bullardi Chero, Cruces, Sáez, Carolina, Camargo, Portes Santos and Luque. Evidence of these three hexabothriid species from South Africa suggests that hexabothriids have more globally widespread distributions, and that they are less host-specific than previously considered. A detailed summary of hexabothriids reported from the triakid hosts, Galeorhinus galeus and Mustelus mustelus is provided, and the host-specificity of hexabothriids is discussed.

Prognostic value of acute National Institutes of Health Stroke Scale Items on disability: a registry study of first-ever stroke in the western part of Sweden.

OBJECTIVES: We aimed to study how the individual items of the National Institutes of Health Stroke Scale (NIHSS) at admission predict functional independence 3 months post-stroke in patients with first-ever stroke. SETTING: This registry-based study used data from two Swedish stroke registers (Riksstroke, the mandatory national quality register for stroke care in Sweden, and Väststroke, a local quality stroke register in Gothenburg). PARTICIPANTS: This study included patients with first-ever acute stroke admitted from November 2014 to August 2018, with available NIHSS at admission and modified Rankin Scale (mRS) at 3-month follow-up. PRIMARY OUTCOME: The primary outcome variable was mRS≤1 (defined as an excellent outcome) at 3-month follow-up. RESULTS: We included 1471 patients, mean age was 72 (± 14.5) years, 48% were female, and 66% had mild strokes (NIHSS≤3). In adjusted binary logistic regression analysis, the NIHSS items impaired right motor arm and leg, and impairment in visual field, reduced the odds of an excellent outcome at 3 months ((OR 0.60 (95% CI 0.37 to 0.98), OR 0.60 (95% CI 0.37 to 0.97), and OR 0.65 (95% CI 0.45 to 0.94)). When exploring the effect size of associations between NIHSS items and mRS≤1 p, orientation, language and right leg motor had the largest yet small association. CONCLUSIONS: Stroke patients with scores on the NIHSS items right motor symptoms or visual field at admission are less likely to have an excellent outcome at 3 months. Clinicians should consider the NIHSS items affected, not only the total NIHSS score, both in treatment guidance and prognostics.

Multimodal individualised intervention to prevent functional decline after stroke: protocol of a randomised controlled trial on long-term follow-up after stroke (LAST-long).

INTRODUCTION: Multimodal interventions have emerged as new approaches to provide more targeted intervention to reduce functional decline after stroke. Still, the evidence is contradictory. The main objective of the Life After Stroke (LAST)-long trial is to investigate if monthly meetings with a stroke coordinator who offers a multimodal approach to long-term follow-up can prevent functional decline after stroke. METHODS AND ANALYSIS: LAST-long is a pragmatic single-blinded, parallel-group randomised controlled trial recruiting participants living in six different municipalities, admitted to four hospitals in Norway. The patients are screened for inclusion and recruited into the trial 3 months after stroke. A total of 300 patients fulfilling the inclusion criteria will be randomised to an intervention group receiving monthly follow-up by a community-based stroke coordinator who identifies the participants' individual risk profile and sets up an action plan based on individual goals, or to a control group receiving standard care. All participants undergo blinded assessments at 6-month, 12-month and 18-month follow-up. Modified Rankin Scale at 18 months is primary outcome. Secondary outcomes are results of blood tests, blood pressure, adherence to secondary prophylaxis, measures of activities of daily living, cognitive function, physical function, physical activity, patient reported outcome measures, caregiver's burden, the use and costs of health services, safety measures and measures of adherence to the intervention. Mixed models will be used to evaluate differences between the intervention and control group for all endpoints across the four time points, with treatment group, time as categorical covariates and their interaction as fixed effects, and patient as random effect. ETHICS AND DISSEMINATION: This trial was approved by the Regional Committee of Medical and Health Research Ethics, REC no. 2018/1809. The main results will be published in international peer-reviewed open access scientific journals and to policy-makers and end users in relevant channels. TRIAL REGISTRATION NUMBER: ClincalTrials.gov Identifier: NCT03859063, registered on 1 March 2019.

Gyrodactylus molweni sp. n. (Monogenea: Gyrodactylidae) from Chelon richardsonii (Smith, 1846) (Mugilidae) from Table Bay, South Africa.

Gyrodactylus molweni sp. n. is described from the body surface and fins of the South African mullet, Chelon richardsonii (Smith, 1846) collected from Table Bay Harbour, Cape Town and is compared to five other Gyrodactylus species described from grey mullets globally namely G. zhukovi Ling, 1963 and G. mugili Zhukov, 1970 from Planiliza haematocheila (Temminck and Schlegel, 1845); G. mugelus Rawson, 1973 from Mugil cephalus L.; G. curemae Conroy and Conroy, 1985 from Mugil curema Valenciennes, 1836 and G. xiamenensis Zang,Yang and Liu, 2001 from Planiliza macrolepis (Smith, 1846). Morphologically, G. molweni sp. n. has prominent ventral bar processes that near cover the hamulus roots, marginal sickles with large rhomboid heels, slender shafts and fine points that extend beyond the sickle toes. Gyrodactylus molweni sp. n. can, however, be readily differentiated: G. mugili and G. xiamenensis have ventral bars with small ventral processes; G. zhukovi has marginal hooks sickles with slender shafts and proportionately short points and open-faced blades; G. mugelus possesses marginal hook sickles with deep, rounded heels, forward slanting shafts and an angular, square line to the inner face of the blades. Although the length of the marginal hooks of G. curemae are similar to G. molweni sp. n., their hamuli are double the size. A GenBank BlastN search with the 931 bp sequence covering ITS1, 5.8S and ITS2 gave no close hits; the nearest species for which sequences are available is G. nipponensis Ogawa and Egusa, 1978 (identity 96.56%, 899/931 bp). The proposal of G. molweni sp. n. as a new species, therefore, is well supported by both the molecular and morphological analyses presented herein. This Gyrodactylus species is the first to be described from C. richardsonii and only the second Gyrodactylus species to be described from the marine environment off the African continent.

Geographical distribution of Gyrodactylus salaris Malmberg, 1957 (Monogenea, Gyrodactylidae).

BACKGROUND: Gyrodactylus salaris Malmberg, 1957 is an OIE (Office International des Epizooties)-listed parasitic pathogen and had until the current study been reported from 19 countries across Europe, although many of these records require confirmation. The last comprehensive evaluation regarding the distribution of G. salaris, however, was made in 2007, although some of the states identified as being G. salaris-positive were ascribed this status based on misidentifications, on partial data resulting from either morphological or molecular tests, or from records that have not been revisited since their early reporting. It is thus important to go through the reports on G. salaris to obtain a status for each country. METHODS: To provide a revised update of the G. salaris distribution, a literature review was necessary. This literature, however, was not always readily accessible and, in certain cases, the article only made superficial reference to the parasite without providing details or data to support the identification. In most cases, the original specimens were not deposited in a national collection. Additional Gyrodactylus material for the current study was obtained from selected salmonid populations with the aim to contribute to current understanding regarding the distribution of G. salaris. Additional parasite material collected for this study was processed following standard procedures for species identification in Gyrodactylus [1]. RESULTS: From the work conducted in the current study, G. salaris is reported from a further three regions in Italy, alongside three other species, and appears to occur extensively throughout central Italy without causing significant mortalities to its rainbow trout, Oncorhynchus mykiss (Walbaum), host. The analysis of archive material from G. salaris-positive farms would suggest that G. salaris has been in this country since at least 2000. Material obtained from rainbow trout from Finland and Germany are confirmed as G. salaris, supporting existing data for these countries. No specimens of G. salaris, however, were found in the additional Gyrodactylus material obtained from rainbow trout reared in Portugal and Spain. A morphologically similar species, Gyrodactylus teuchis Lautraite, Blanc, Thiery, Daniel et Vigneulle, 1999, however, was found. CONCLUSIONS: Following the present review, Gyrodactylus salaris is reported from 23 out of 50 recognised states throughout Europe; only records from 14 of these states have been confirmed by either morphology and/or by an appropriate molecular test and are considered valid, while only nine of these records have been confirmed by a combination of both methods.

Multi-agent in situ hybridization confirms Ca. Branchiomonas cysticola as a major contributor in complex gill disease in Atlantic salmon.

Gill diseases may cause high mortalities in farmed Atlantic salmon. In seawater reared fish co-infections involving the epitheliocystis associated bacterium Ca. Branchiomonas cysticola, the microsporidian Desmozoon lepeophtherii, the causative agent of amoebic gill disease Paramoeba perurans and salmon gill poxvirus are common and histopathological lesions may be complex. Here, we report detection of these agents utilising multiplex real-time PCR and link the presence of agents to histopathologically visible gill lesions by in situ hybridisation (ISH) utilising RNAscope®. We show that Ca. Branchiomonas cysticola infections may remain undetected if diagnostic investigations are restricted to histopathology alone. Further, positive in situ labelling of Ca. Branchiomonas cysticola was observed within epitheliocysts, but also in small foci within areas of inflammation and necrosis in which histologically detectable epitheliocysts were not visible. In situ labelling of D. lepeophtherii corresponded well with tissue distribution patterns previously associated with this microsporidian. Salmon gill poxvirus was associated with apoptotic gill epithelial cells, while Ca. Piscichlamydia salmonis could not be associated with pathological changes. The multiplex real-time PCRs utilised were rapid and sensitive diagnostic tools and the results corresponded well with ISH. This study shows that the agents involved in complex gill disease can be linked to lesions using ISH and suggests that Ca. B. cysticola plays a crucial role in the development of gill disease in the farming of salmon in Norway.