Tropical Medicine and Infectious Disease

Brief Report Brief Report e-DiagnosisE-Diagnosis inin MedicalMedical ParasitologyParasitology

Harsha Sheorey Medical Microbiologist, St Vincent’sVincent’s Hospital,Hospital, MelbourneMelbourne 3065,3065, Australia;Australia; [email protected]@svha.org.au Received: 19 December 2019; Accepted: 1 January 2020; Published: 3 January 2020  Received: 19 December 2019; Accepted: 1 January 2020; Published: 3 January 2020  Abstract: Over the past decade or two, the teaching of laboratory diagnostic has been Abstract: Over the past decade or two, the teaching of laboratory diagnostic parasitology has been neglected in Australasia, as parasitic infections are relatively uncommon. As a consequence, neglected in Australasia, as parasitic infections are relatively uncommon. As a consequence, expertise expertise in medical parasitology is dwindling. A team of international experts (including Professor in medical parasitology is dwindling. A team of international experts (including Professor John John Goldsmid) has been formed to help in the diagnosis of human parasitic infections. The team Goldsmid) has been formed to help in the diagnosis of human parasitic infections. The team includes includes experts from Australia, Europe, South Africa and the USA. Some senior members of the experts from Australia, Europe, South Africa and the USA. Some senior members of the team are team are excellent morphologists, and we have both human and veterinary parasitologists who help excellent morphologists, and we have both human and veterinary parasitologists who help with with molecular diagnosis in difficult cases. molecular diagnosis in difficult cases. Keywords: e-Diagnosis; morphologist; molecular parasitology Keywords: e-Diagnosis; morphologist; molecular parasitology

Professor John Goldsmid has been a doyen of medical parasitology in Australia for many years. I was Professor John Goldsmid has been a doyen of medical parasitology in Australia for many years. honored to meet John during our inaugural parasitology master class organized by the Australian I was honored to meet John during our inaugural parasitology master class organized by the Society for Microbiology (ASM) in Hobart in 2009 (Figure1). Goldsmid’s knowledge, dedication and Australian Society for Microbiology (ASM) in Hobart in 2009 (Figure 1). Goldsmid’s knowledge, teaching in this field must be carried on for the future generations of health professionals. His attitude dedication and teaching in this field must be carried on for the future generations of health towards learning is exemplary (see response to Case 4 below). professionals. His attitude towards learning is exemplary (see response to Case 4 below).

Figure 1. The inaugural A Australianustralian parasitology master class.class.

Trop. Med. Infect. Dis. 2020, 5, x; doi: FOR www.mdpi.com/journal/tropicalmed Trop. Med. Infect. Dis. 2020, 5, 8; doi:10.3390/tropicalmed5010008 www.mdpi.com/journal/tropicalmed Trop. Med. Infect. Dis. 2020, 5, 8 2 of 7

Trop. Med. Infect. Dis. 2020, 5, x FOR PEER REVIEW 2 of 7 Over the past decade or two, the teaching of laboratory diagnostic parasitology has been neglected in AustralasiaOver the because past decade parasitic or infections two, the teaching are relatively of laboratory uncommon. diagnostic As a consequence, parasitology the has expertise been inneglected medical parasitologyin Australasia is because dwindling. parasitic While infections on a sabbaticalare relatively at theuncommon. Centre for As Diseasea consequence Control, the and expertise in medical parasitology is dwindling. While on a sabbatical at the Centre for Disease Control Prevention in Atlanta (CDC), USA, I came across a method of diagnosing parasites that the parasitology and Prevention in Atlanta (CDC), USA, I came across a method of diagnosing parasites that the department there had adopted to help less experienced health professionals. A picture of an unknown parasitology department there had adopted to help less experienced health professionals. A picture malarial parasite from Africa was sent to the parasitologists on a smart phone, and, after looking at the of an unknown malarial parasite from Africa was sent to the parasitologists on a smart phone, and, image, they were able to send back a diagnosis of malarial parasite to a species level. This method after looking at the image, they were able to send back a diagnosis of malarial parasite to a species couldlevel. easily This bemethod adopted could in Australia,easily be adopted I thought. in Australia, Thus, a team I thought. of experts Thus who, a team have of helped experts in who diagnosing have andhelped managing in diagnosing parasitic and infections managing was parasitic put together. infections was put together. 1. Why e-Diagnosis? 1. Why e-Diagnosis? Less importance/attention is given to parasitic diseases in humans in the developed world • • Less importance/attention is given to parasitic diseases in humans in the developed world (including(including Australia). Australia). • The re-distribution of parasites due to human/ movement and environmental and practice • The re-distribution of parasites due to human/animal movement and environmental and changespractice has changes brought has parasites brought parasites into the developedinto the developed world. world. • ThereThere are are declining declining numbers numbers of ofexperts experts inin thisthis field,field, especially morphologists morphologists.. • • TheThe internet internet ‘savvy’ ‘savvy public’ public who who ‘self-diagnose’‘self-diagnose’ want to to ‘ ‘confirm’confirm’ their their thoughts thoughts.. • 2.2. Who Who Is Is the the Team? Team? TheThe team team includes includes parasitologists parasitologists andand other experts from from Australia, Australia, Europe, Europe, South South Africa Africa,, and and thethe USA. USA. Some Some senior senior members members ofof thethe teamteam are excellent morphologists morphologists,, and and we we have have both both human human andand veterinary veterinary parasitologists parasitologists whowho help with with molecular molecular diagnosis diagnosis (Fig (Figureure 2).2 ).The The author author acts acts as a as coordinator of the group. The team has helped diagnose a few unusual parasites, leading to a coordinator of the group. The team has helped diagnose a few unusual parasites, leading to publications [1–4]. publications [1–4].

FigureFigure 2.2. The way e e-Diagnosis-Diagnosis works. works. 3. What Kind of Material/Advice is Sought? 3. What Kind of Material/Advice is Sought? Material: Material: • ImagesImages for for identification identification (macroscopic (macroscopic andand microscopic)microscopic) • • HistologyHistology sections sections of of tissues tissues with with possiblepossible parasites • Advice: • Advice:Molecular identification/availability • Interpretation/availability of serology for parasites Molecular identification/availability • • Advice on testing to exclude parasites Interpretation/availability of serology for parasites •

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Advice on testing to exclude parasites • Management of parasitic infections (treatment and follow-up)—OzBug (Australian email group • Trop.of Med. interested Infect. Dis. 2020 clinical, 5, x FOR experts) PEER REVIEW 3 of 7 Advice on/to “internet-savvy” patient concerns • • Management of parasitic infections (treatment and follow-up)—OzBug (Australian email group 4. Typeof ofinterested Parasites clinical Received experts) from Intestine, Tissues or Blood • Advice on/to “internet-savvy” patient concerns Protozoa • 4. TypeNematodes of Parasites Received from Intestine, Tissues or Blood • • TrematodesProtozoa • • CestodesNematodes • • Trematodes • • Cestodes Pseudo-parasites and artefacts • • Arthropods • Pseudo-parasites and artefacts 5. Cases 5. Cases 5.1. Case 1: Lump in the Back (Melbourne, VIC, Australia) 5.1. Case 1: Lump in the Back (Melbourne, Vic) A lump in the back (right scapular region) (Figure3A) in a young women was biopsied to rule out sarcoma.A lump in Its the histology back (right showed scapular a region) helminth (Figure with 3A) morphological in a young women features was biopsied suggestive to rule of a filarial worm—out sarcoma. Its histologyor Dirofilaria showedspecies a helminth (Figure with3 B).morphological This prompted features the suggestive aid of an infectiousof a filarial diseases consult,worm—Onchocerca and history-taking or revealed species that(Figure the 3B). patient This prompted was a veterinarian the aid of an with infectious extensive diseases exposure to consult, and history-taking revealed that the patient was a veterinarian with extensive exposure to various both in Australia and Africa. Apart from swelling, the patient felt ‘movement’ inside various animals both in Australia and Africa. Apart from swelling, the patient felt ‘movement’ inside their nodule. A lumpectomy was performed, and, in the laboratory, the worm fell out of the lump their nodule. A lumpectomy was performed, and, in the laboratory, the worm fell out of the lump (Figure(Figure3 C).3C). On On examination,examination, this this was was confirmed confirmed to be to a be filarial a filarial worm worm with features with features of Onchocerca of Onchocerca speciesspecies (cuticular (cuticular rings and and subcuticular subcuticular striae) striae) (Figure (Figure 3D),3 mostD), most likely likely a zoonotic a zoonotic OnchocercaOnchocerca lupi or lupi or OnchocercaOnchocercavolvulus volvulus.

(A) (B)

(C) (D)

FigureFigure 3.3. ((A) PETPET/CT/CT scan scan of of back back showing showing a lump a lump in inthe the back back (red (red arrow). arrow). (B) Histology (B) Histology section section of of the biopsythe biopsy showing showing a a nematode helminth helminthwith with featuresfeatures of of a a filarial filarial worm. worm. (C) ( CIntertwined) Intertwined worm(s) worm(s) fell outfellof out the of surgicallythe surgically removed removed lump. lump. (D (D) On) On exam, exam, thesethese werewere found found to to be be two, two, a athicker thicker female female and a and a thinner male. thinner male.

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Trop. Med. Infect. Dis. 2020, 5, x FOR PEER REVIEW 4 of 7 e-Diagnosis members views: e-DiagnosisInitial histology: members Expertsviews: were convinced the section had features of a filarial helminth (ridges and internalInitial histology: structures), Experts most likelywere aconvinced zoonotic Onchocercathe section specieshad features or Dirofilaria of a filarialspecies. helminth (ridges and internalAfter the stru wormctures), was most extracted, likely experts a zoonotic were Onchocerca convinced species that the or morphology Dirofilaria species. was most consistent with OnchocercaAfter the wormspecies. was A extracted, PCR-based experts sequencing were convinced method confirmed that the morphology it to be O. volvulus was most(1). consistent with Onchocerca species. A PCR-based sequencing method confirmed it to be O. volvulus (1). 5.2. Case 2: Lump in the Axilla (Brisbane, QLD, Australia) 5.2. Case 2: Lump in the Axilla (Brisbane, Qld) A 23 year old from Brisbane, Australia, presented with an axillary mass for six months. He noticed movementA 23 year within, old and from a small Brisbane, worm Australia was expressed., presented This with was an placed axillary in formalin. mass for Thesix dimensionsmonths. He werenoticed 1.2 mmmovement wide by within 3.85 mm, and long a small (Figure worm4). No was relevant expressed. travel history This was was placed noted. in formalin. The dimensions were 1.2 mm wide by 3.85 mm long (Figure 4). No relevant travel history was noted.

(A) (B)

FigureFigure 4.4.( A(A)) Picture Picture of of histological histological section section of ‘worm’ of ‘worm sent’ tosent the to e-Diagnosis the e-Diagnosis team. (team.B) The (B same) The picture same thatpicture was that labelled was andlabelled sent back and to sent the back team byto Professorthe team Anderson, by Professor showing Anderson various, showing morphological various featuresmorphological that are features consistent that with are anconsistent endopterygota with an larva.endopterygota larva. e-Diagnosis member view: e-Diagnosis member view: ProfessorProfessor EmeritusEmeritus DonDon AndersonAnderson (previously(previously ChallisChallis ProfessorProfessor ofof ZoologyZoology atat SydneySydney University) University) ‘An‘An endopterygotaendopterygota larva,larva, mostmost probablyprobably aa beetlebeetle larvalarva withwith hookedhooked mandibles.mandibles. A lotlot ofof beetlebeetle larvaelarvae burrowburrow into into organic organic substrates. substrates. It’s It’ as pitya pity histopathologists histopathologists almost almost invariably invariably section section things things that arethat more are more easily easily identified identified intact.’ intact.’

5.3. Case 3: Larvae, Adults and Eggs in CSF/Brain (Adelaide, SA, Australia) 5.3. Case 3: Larvae, Adults and Eggs in CSF/Brain (Adelaide, SA) A 73 year old woman was admitted to a regional South Australia hospital (past history with A 73 year old woman was admitted to a regional South Australia hospital (past history with unspecifiedunspecified abdominal abdominal pain). pain). The The patient patient was on was low on dose low methotrexate dose methotrexate and etanercept and foretanercept Rheumatoid for arthritisRheumatoid (RA). arthritis She was (RA) transferred. She was to Royal transferred Adelaide to Hospital Royal Adelaide with deteriorating Hospital withneurological deteriorating signs. Theneurological Cerebro-spinal signs. The fluid Cerebro (CSF)-spinal taken fluid showed (CSF) 280 taken polymorphs, showed 280 18 polymorphs, monocytes, no18 monocytes, bacteria seen, no Cryptococcal highbacteria protein, seen, and high virology protein and, and bacterial virology PCR and and bacterial PCR andantigen Cryptococcal all negative. The all CSF neg takenative. twoThe daysCSF later taken showed two days 2500 later polymorphs, showed 342500 monocytes, polymorphs, no bacteria 34 monocytes, seen; Diff-Quik no b (modifiedacteria seen; Romanowsky Diff-Quik Acanthamoeba Stain)(modified showed Romanowsky “few eosinophils,” Stain) showed and no trophozoites.“few eosinophils,” and nocultures trophozoites. were set Acanthamoeba up. Patient diedcultures less were than set a week up. Patient after admission, died less than and a aweek post after mortem admissio examinationn, and a waspost conducted.mortem examination The CSF atwas post conducted. mortem showedThe CSF 1700at post polymorphs mortem showed (PMN) 1700 (few polymorphs eosinophils), (PMN) a very (few high eosinophils), protein level, a very and Strongyloides Angiostrongylus numeroushigh protein helminth level, larvaeand numerous and eggs helminth(Figure5A,B). larvae and eggs (Fandigure 5 A,B). Strongyloidesserology wasand Angiostrongylus serology was negative. A brain biopsy demonstrated the presence of a helminth. PCR-based sequencing confirmed it to be Halicephalobus gingivalis (Figure 5C).

Trop. Med. Infect. Dis. 2020, 5, 8 5 of 7 negative. A brain biopsy demonstrated the presence of a helminth. PCR-based sequencing confirmed it to be Halicephalobus gingivalis (Figure5C). Trop. Med. Infect. Dis. 2020, 5, x FOR PEER REVIEW 5 of 7

(A) (B) (C)

Figure 5. ((AA)) E Egggg of of parasite parasite in in CSF CSF resembling resembling the the TrichostrongylusTrichostrongylus speciesspecies (~50 (~50 × 2020 µm)µm).. (B) ( Blarva) larva of × parasiteof parasite in inCSF CSF resembling resembling StrongyloidesStrongyloides speciesspecies (~250 (~250 µm).µm). (C (C) )Post Post-mortem-mortem brain brain tissue showing adult worms, larvae and eggs of the helminth (details and additional pictures inin ReferenceReference 2).2). e-Diagnosis members views: e-Diagnosis members views: What fol followslows is a summary of the email discussion that occurred after images were sent to the team. The The experts experts’’ names are in brackets.

• The structurestructure ofof the the esophagus esophagus and and ratio ratio to gutto gut may may also also help help us. Theus. twoThe illustratedtwo illustrated larvae larvae could • justcould be just rhabditiform be rhabditiform and hence and nohenc notchinge no notching of tail; thereof tail does; there seem does to seem be a darker to be a region darker near region the genitalnear the primordium genital primordium zone in onezone photo, in one but photo we, also but lackwe also ability lack to ability gain more to gain definition more definition of these keyof these points. key (Norbertpoints. (Norbert Ryan). Ryan). • The egg lookslooks veryvery muchmuch like like a aTrichostrongylus Trichostrongylusegg egg (but (but the the size size of theof the eggs eggs is too is too small), small), and and the • larvaethe larvae have have features features that that are reminiscent are reminiscent of that of speciesthat species also, particularlyalso, particularly the wavy the wavy pattern pattern of the intestineof the intestine (John Walker).(John Walker). • First of all, very few documented cases (of Strongyloides) in the brain/CSF. Have no explanation First of all, very few documented cases (of Strongyloides) in the brain/CSF. Have no explanation for • for eggs/rhabditiform larvae in this site (Lynne Garcia). eggs/rhabditiform larvae in this site (Lynne Garcia). • I don’t think it is the usual suspects, namely Strongyloides or Angiostyrongylus, on morphological I don’t think it is the usual suspects, namely Strongyloides or Angiostyrongylus, on morphological • grounds. Eggs are not normally seen in . They look most like stages of grounds. Eggs are not normally seen in angiostrongyliasis. They look most like stages of , but that infection does not disseminate, or at least there are no reports thereof. Trichostrongylus, but that infection does not disseminate, or at least there are no reports thereof. Thus, I think this is a very unusual aberrant infection with Trichostrongylus or similar nematode Thus, I think this is a very unusual aberrant infection with Trichostrongylus or similar nematode that is not normally found in humans (John Frean). that is not normally found in humans (John Frean). • I found a lovely description of all stages of Halicephalobus gingivalis in Anderson et al. (1998). This I found a lovely description of all stages of Halicephalobus gingivalis in Anderson et al. (1998). • is now regarded as the correct name for the parasite. The drawings are very consistent with what This is now regarded as the correct name for the parasite. The drawings are very consistent with is in the images. Note the elongated eggs, the recurved ovaries, and of course the rhabditoid what is in the images. Note the elongated eggs, the recurved ovaries, and of course the rhabditoid esophagus (Rick Speare). esophagus (Rick Speare). • Please don’t waste the you have by rushing in and using them immediately for PCR. Please don’t waste the nematodes you have by rushing in and using them immediately for PCR. • They should be morphologically described first; then do PCR. Additionally, some should be kept Theyin case should it is a new be morphologically species. Don’t forget described the light first; microscopy then do PCR. and measurements Additionally,, some as well should as SEM. be keptOf course in case, the it is latter a new looks species. more Don’t spectacular forget the but light usually microscopy carries less and taxonomic measurements, weight as (Rick well asSpeare) SEM.. Of course, the latter looks more spectacular but usually carries less taxonomic weight • (RickThe sequence Speare). results have come back, and the worms are positive for Halicephalobus gingivalis. The sequence results have come back, and the worms are positive for Halicephalobus gingivalis. • The 28s D2/D3 primers from Nadler’s 2003 paper were used and got the sequence found below The(2). This 28s D2 sample/D3 primers was 99% from identical Nadler’s to 2003GenBank paper accession were used AY294177 and got the over sequence 786 base found pairs below (Anson (2). ThisKohler) sample. was 99% identical to GenBank accession AY294177 over 786 base pairs (Anson Kohler). • The combination of case details, plus description of the nematodes, plus PCR gives the maximum • The combination of case details, plus description of the nematodes, plus PCR gives the value.maximum Fabulous value. e ffFabulousort! Good effort! example Good of theexample value of athe collaborative value of a collaborative dispersed group dispersed in diagnosing group unusualin diagnosing parasites! unusual (Rick parasites! Speare) (Rick Speare)

5.4. Case 4: Worm Passed in Feces (Wollongong, NSW) From Dr. Peter Newton, Pathologist in Wollongong: This worm was sent to us for identification from one of our regional hospitals 80 km away. The history that we received from the ID physician who sent it to us was that of a 75 year old female patient admitted with severe Clostridium difficile- associated disease (CDAD). Her condition has apparently been slowly improving. Yesterday, she was

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5.4. Case 4: Worm Passed in Feces (Wollongong, NSW, Australia) From Dr. Peter Newton, Pathologist in Wollongong: This worm was sent to us for identification from Trop.one Med. of our Infect. regional Dis. 2020 hospitals, 5, x FOR PEER 80 km REVIEW away. The history that we received from the ID physician who6 of sent 7 it to us was that of a 75 year old female patient admitted with severe Clostridium difficile-associated incontinentdisease (CDAD). of feces Her on condition the ward hasfloor apparently, and this worm been slowly was seen improving. wriggling Yesterday, out of her she feces was on incontinent the floor (Fofigure feces 6) on. the ward floor, and this worm was seen wriggling out of her feces on the floor (Figure6). BasedBased on on a a Google Google search search,, we we think think that that it it is is a a “hammerhead “hammerhead worm” worm” of of the the BipaliumBipalium genus.genus. We We assumeassume it it was was on on the the floor floor and crawled into her feces,feces, oror thethe fecesfeces landedlandedon on the the worm worm on on the the floor. floor. It Itwas was a a rather rather wet wet morning morning on on the the New New South South Wales Wales (NSW) (NSW) coast coast yesterday, yesterday so, presumably so presumably came came onto ontothe wardthe ward on someone’s on someone shoe.’s shoe. Would Would value value your your opinion. opinion.

Figure 6. Worm thought to have wriggled out of feces on the floor in an incontinent patient. Figure 6. Worm thought to have wriggled out of feces on the floor in an incontinent patient. ee-Diagnosis-Diagnosis members members view/comments: view/comments: DrDr.. John John Walker Walker (ex (ex-Head-Head of of Parasitology, Parasitology, Westmead Westmead Hospital, Hospital, Sydney) Sydney) TheThe worm worm is is BipaliumBipalium kewense kewense, ,first first described described at at Kew Kew Gardens Gardens but but originally originally from from Southeast Southeast Asia. Asia. NowNow has a a cosmopolitan cosmopolitan distribution distribution as a as consequence a consequence of trade of in trade botanical in botanical specimens. specimens. It’s a predatory, It’s a pfree-livingredatory, free planarian.-living planarian. ProfProf John John Goldsmid Goldsmid (UTas, (UTas, Hobart) Hobart) MyMy goodness goodness—we—we live live and and learn, learn, even even in in retirement!! retirement!! I Ihave have never never seen seen this this worm worm or or heard heard of of a a ““hammerheadhammerhead worm worm.”.” Peter Peter’s’s tentative tentative identification identification seems seems reasonable reasonable and and probable. probable. I Icongratul congratulateate himhim on on his his detective detective work work..

6. Limitations of This Diagnostic Procedure 6. Limitations of This Diagnostic Procedure Poor images may be sent: •• Poor images may be sent: – Not enough images: multiple images at different magnification levels and that cover key - Not enough images: multiple images at different magnification levels and that cover key parts of the parasite are not always sent. parts of the parasite are not always sent. – Poor quality photos that are out of focus or focused on wrong part may be sent. The new - Poor quality photos that are out of focus or focused on wrong part may be sent. The new smart phones take very good images, so this should not be an excuse. smart phones take very good images, so this should not be an excuse. • No relevant history: Age, , travel, exotic food, contact with animals, medication, immuneNo relevant status history: are all Age,important epidemiology,, and, quite travel, often,exotic this is food,not communicated. contact with animals, medication, • • Arthropodsimmune status or large are all worms important, received and, in quite formalin often, and this sectioned: is not communicated. Arthropods and large worms areArthropods better photographed or large worms whole received and not in sectioned. formalin When and sectioned: stored in Arthropods70% alcohol, and formalin large makes worms • specimenare betters photographed‘brittle.’ whole and not sectioned. When stored in 70% alcohol, formalin makes • ‘Degeneratespecimens ‘brittle.’’ or dying parasite (histology-section): Morphology may not be typical enough to make a diagnosis. • Not enough follow up material: The specimen or multiple images should be held onto if required for further identification. • Direct contact with non-scientific public (via social media and internet): – Abuse: Some members of the public do not like answers that they do not want to hear and can get abusive.

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‘Degenerate’ or dying parasite (histology-section): Morphology may not be typical enough to • make a diagnosis. Not enough follow up material: The specimen or multiple images should be held onto if required • for further identification. Direct contact with non-scientific public (via social media and internet): • - Abuse: Some members of the public do not like answers that they do not want to hear and can get abusive. - Legal aspects: Advice/opinions given over limited information and presented this way should be taken in the context of clinical pictures. It should be made clear that this is an opinion, and a clinical judgment should be made by the treating doctor. Advice on treatment (drugs) should be avoided over internet/social media, as other medical history information needs to be taken in consideration. Wrong interpretation/understanding can lead to legal ramifications.

7. Summary and the Future Given the decreasing expertise in medical parasitology, this type of diagnostic help will become common and convenient. A team of experts can be put together from various parts of the world, and images and questions can be shared over electronic media. Additionally, sending bits of specimens by post to experts anywhere in the world is now possible in case molecular techniques are required for diagnosis. However, the legal aspects of giving advice and possible abuse over electronic or social media should be kept in mind.

Conflicts of Interest: The author declares no conflict of interest.

References

1. Crowe, A.; Koehler, A.V.; Sheorey, H.; Tolpinrud, A.; Gasser, R.B. PCR-coupled sequencing achieves specific diagnosis of in a challenging clinical case, to underpin effective treatment and clinical management. Infect. Genet. Evol. 2018, 66, 192–194. [CrossRef][PubMed] 2. Lim, C.K.; Crawford, A.; Moore, C.V.; Gasser, R.B.; Nelson, R.; Koehler, A.V.; Bradbury, R.S.; Speare, R.; Dhatrak, D.; Weldhagen, G.F. First human case of fatal Halicephalobus gingivalis meningoencephalitis in Australia. J. Clin. Microbiol. 2015, 53, 1768–1774. [CrossRef][PubMed] 3. McKelvie, P.; Reardon, K.; Bond, K.; Spratt, D.M.; Gangell, A.; Zochling, J.; Daffy, J. A further patient with parasitic myositis due to Haycocknema perplexum, a rare entity. J. Clin. Neurosci. 2013, 20, 1019–1022. [CrossRef][PubMed] 4. Koehler, A.V.; Spratt, D.M.; Norton, R.; Warren, S.; Mcewan, B.; Urkude, R.; Murth, S.; Robertson, T.; Mccallum, N.; Parsonson, F.; et al. More parasitic myositis cases in humans in Australia, and the definition of genetic markers for the causative agents as a basis for molecular diagnosis. Infect. Genet. Evol. 2016, 44, 69–75. [CrossRef][PubMed]

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