Practical Guide to Diagnostic Parasitology E-Book

Lynne Shore Garcia

• Herausgeber: John Wiley & Sons
• Kategorie: Wissenschaft und neue Technologien
• Serie: ASM
• Sprache: Englisch

Seitenzahl: 1199

Veröffentlichungsjahr: 2021

Beschreibung

In the 21st century the field of diagnostic medical parasitology continues to see dramatic changes, including newly recognized pathogens and the changing endemicity and classification of familiar organisms; neglected tropical diseases and the impact of global climate change; and new methodologies and risk management issues. This classic clinical laboratory parasitology reference, now in its third edition, has been extensively revised and updated in a new full-color format. Still organized to provide maximum help to the user, particularly from the bench perspective, every section has been expanded with new images and discussion.

Specimen collection, preservation, and testing options are thoroughly discussed, from the routine ova and parasite examination to blood films, fecal immunoassays, and the newer molecular test panels. Specific test procedures, laboratory methods and reagents, and algorithms are provided. The ever-helpful "FAQ" section of commonly asked questions now offers expanded information on stool specimen fixatives and testing, thorough coverage of new techniques, and advice on reporting and commenting on results.

The heart of the Guide, covering identification of individual pathogens, has been expanded with more discussion and comparison of organisms and dozens of new color images. An entirely new section has been added that uses extensive figures and new tables to illustrate common problems with differentiating organisms from one another and from possible microscopic artifacts. The final section has been reorganized to include identification keys and dozens of tables summarizing organism characteristics to assist the bench microbiologist with routine diagnostic testing methods.

If you are looking for online access to the latest clinical microbiology content, please visit www.wiley.com/learn/clinmicronow.

Leseprobe

Table of Contents

Cover

Title Page

Copyright

Dedication

Preface

ACKNOWLEDGMENTS

About the Author

SECTION 1: Philosophy and Approach to Diagnostic Parasitology

Neglected Tropical Diseases

Why Perform Diagnostic Parasitology Testing?

Travel

Population Movements

Control Issues

Climate Change

Epidemiologic Considerations

Compromised Patients; Potential Sex Bias Regarding Infection Susceptibility; Aging

Approach to Therapy

Who Should Perform Diagnostic Parasitology Testing?

Laboratory Personnel

Nonlaboratory Personnel

Where Should Diagnostic Parasitology Testing Be Performed?

Inpatient Setting

Outpatient or Referral Setting

Decentralized Testing

Physician Office Laboratories

Over‐the‐Counter (Home Care) Testing

Field Sites

What Factors Should Precipitate Testing?

Travel and Residence History

Immune Status of the Patient

Clinical Symptoms

Documented Previous Infection

Contact with Infected Individuals

Potential Outbreak Testing

Occupational Testing

Therapeutic Failure

What Testing Should Be Performed?

Routine Tests

Special Testing and Reference Laboratories

Specialized Referral Test Options—DPDx and Other Sites

Other (Nonmicrobiological) Testing

What Factors Should Be Considered in Development of Test Menus?

Physical Plant

Client Base

Customer Requirements and Perceived Levels of Service

Personnel Availability and Level of Expertise

Equipment

Budget

Risk Management Issues Associated with Stat Testing

Primary Amebic Meningoencephalitis

Granulomatous Amebic Encephalitis and Amebic Keratitis

Request for Blood Films

Automated Instrumentation

Patient Information

Conventional Microscopy

Table 1.1 Common features of the neglected tropical diseases

SECTION 2: Parasite Classification and Relevant Body Sites

Protozoa (Intestinal)

Amebae, Stramenopiles

Flagellates

Ciliates

Apicomplexa (Including Coccidia)

Microsporidia (Now Classified with the Fungi)

Protozoa (Other Body Sites)

Amebae

Flagellates

Apicomplexa (Including Coccidia)

Microsporidia (Now Classified with the Fungi)

Protozoa (Blood and Tissue)

Apicomplexa (Including Sporozoa)

Flagellates

Nematodes (Intestinal)

Nematodes (Tissue)

Nematodes (Blood and Tissue)

Cestodes (Intestinal)

Cestodes (Tissue)

Trematodes (Intestinal)

Trematodes (Liver and Lungs)

Trematodes (Blood)

Pentastomids

Acanthocephala

Table 2.1 Classification of human parasites

Table 2.2 Cosmopolitan distribution of common parasitic infections

Table 2.3 Body sites and possible parasites recovered

SECTION 3: Collection Options

Safety

Collection of Fresh Stool Specimens

Collection Method

Number of Specimens To Be Collected

Collection Times

Posttherapy Collection

Specimen Type, Stability, and Need for Preservation

Preservation of Stool Specimens

Overview of Preservatives

Formalin

Sodium Acetate‐Acetic Acid‐Formalin (SAF)

Schaudinn's Fluid

Schaudinn's Fluid Containing PVA (Mercury Base)

Schaudinn's Fluid Containing PVA (Copper Base, Zinc Base)

Single‐Vial Collection Systems (Other than SAF)

Universal Fixative (Total‐Fix)

Quality Control for Preservatives

Procedure Notes for Use of Preservatives (Stool Fixative Collection Vials)

Procedure Limitations for Use of Preservatives (Stool Fixative Collection Vials)

Collection of Blood

Collection and Processing

Stat Test Requests and Risk Management Issues

Collection of Specimens from Other Body Sites

Table 3.1 Fecal specimens for parasites: options for collection and processing

Table 3.2 Approaches to stool parasitology: test ordering

Table 3.3 Preservatives and procedures commonly used in diagnostic parasitology (stool specimens)

Table 3.4 Advantages of thin and thick blood films

Table 3.5 Advantages and disadvantages of buffy coat films

Table 3.6 Potential problems of using EDTA anticoagulant for the preparation of thin and thick blood films

Table 3.7 Body sites and possible parasites recovered (trophozoites, cysts, oocysts, spores, adults, larvae, eggs, amastigotes, and trypomastigotes)

SECTION 4: Specimen Test Options: Routine Diagnostic Methods and Body Sites

Ova and Parasite Examination of Stool Specimens

Other Diagnostic Methods for Stool Specimens

Culture of Larval‐Stage Nematodes

Estimation of Worm Burdens through Egg Counts

Hatching Test for Schistosome Eggs

Screening Stool Samples for Recovery of a Tapeworm Scolex

Testing of Other Intestinal Tract Specimens

Examination for Pinworm

Sigmoidoscopy Material

Duodenal Drainage Material

Duodenal Capsule Technique (Entero‐Test)

Urogenital Tract Specimens

Sputum

Aspirates

Biopsy Specimens

Blood

Thin Blood Films

Thick Blood Films

Blood Staining Methods

Buffy Coat Films

QBC Microhematocrit Centrifugation Method

Knott Concentration

Membrane Filtration Technique

Culture Methods

Animal Inoculation and Xenodiagnosis

Antibody and Antigen Detection

Antibody Detection

Antigen Detection, Nucleic Acid‐Based Tests, and Molecular Panels

Intradermal Tests

UV Autofluorescence

Table 4.1 Body sites, procedures and specimens, recommended methods and relevant parasites, and comments

Table 4.2 Serologic, antigen, and probe tests used in the diagnosis of parasitic infections

SECTION 5: Specific Test Procedures and Algorithms

Microscopy

CALIBRATION OF THE MICROSCOPE

Ova and Parasite Examination

DIRECT WET FECAL SMEAR

CONCENTRATION (Sedimentation and Flotation)

SEDIMENTATION CONCENTRATION (Formalin‐Ethyl Acetate)

SEDIMENTATION CONCENTRATION USING THE UNIVERSAL FIXATIVE (Total‐Fix)

FLOTATION CONCENTRATION (Zinc Sulfate)

PERMANENT STAINED SMEAR

Stains Used in the Permanent Stained Smear

TRICHROME STAIN (Wheatley's Method)

IRON HEMATOXYLIN STAIN (Spencer‐Monroe Method)

IRON HEMATOXYLIN STAIN (Tompkins‐Miller Method)

MODIFIED IRON HEMATOXYLIN STAIN (Incorporating the Carbol Fuchsin Step)

CHLORAZOL BLACK E STAIN

Specialized Stains for Coccidia and Microsporidia

KINYOUN'S ACID‐FAST STAIN (Cold Method)

MODIFIED ZIEHL‐NEELSEN ACID‐FAST STAIN (Hot Method)

CARBOL FUCHSIN NEGATIVE STAIN FOR CRYPTOSPORIDIUM (W. L. Current)

RAPID SAFRANIN METHOD FOR CRYPTOSPORIDIUM (D. Baxby)

RAPID SAFRANIN METHOD FOR CYCLOSPORA, USING A MICROWAVE OVEN (Govinda Visvesvara)

AURAMINE O STAIN FOR APICOMPLEXA (INCLUDING COCCIDIA) (Thomas Hänscheid)

MODIFIED TRICHROME STAIN FOR MICROSPORIDIA (Weber, Green Counterstain)

MODIFIED TRICHROME STAIN FOR MICROSPORIDIA (Ryan, Blue Counterstain)

MODIFIED TRICHROME STAIN FOR MICROSPORIDIA (Evelyn Kokoskin, Hot Method)

Fecal Immunoassays for Intestinal Protozoa

ENTAMOEBA HISTOLYTICA

CRYPTOSPORIDIUM SPP.

GIARDIA LAMBLIA

KITS UNDER DEVELOPMENT

COMMENTS ON THE PERFORMANCE OF FECAL IMMUNOASSAYS

ENZYME IMMUNOASSAYS (Antigen Detection, No Centrifugation Recommended)

FLUORESCENCE (Visual Identification of the Organisms, Centrifugation Recommended)

LATERAL‐FLOW CARTRIDGES (Antigen Detection, No Centrifugation Recommended)

Larval Nematode Culture

HARADA‐MORI FILTER PAPER STRIP CULTURE

BAERMANN CONCENTRATION

AGAR PLATE CULTURE FOR STRONGYLOIDES STERCORALIS

Other Methods for Gastrointestinal Tract Specimens

EXAMINATION FOR PINWORM (Cellulose Tape Preparations)

SIGMOIDOSCOPY SPECIMENS (Direct Wet Smear)

SIGMOIDOSCOPY SPECIMENS (Permanent Stained Smear)

DUODENAL ASPIRATES

Methods for Urogenital Tract Specimens

RECEIPT OF DRY SMEARS

DIRECT SALINE MOUNT

PERMANENT STAINED SMEAR

URINE CONCENTRATION (Centrifugation)

URINE CONCENTRATION (Nuclepore Membrane Filter)

Preparation of Blood Films

THIN BLOOD FILMS

THICK BLOOD FILMS

COMBINATION THICK‐THIN BLOOD FILMS

RISK MANAGEMENT ISSUES ASSOCIATED WITH BLOOD FILMS

USE OF A REFERENCE LABORATORY FOR PARASITE BLOOD DIAGNOSTIC TESTING

BLOOD FILM REPORTING WITH ADDITIONAL REPORT COMMENTS

BUFFY COAT BLOOD FILMS

Blood Stains

STAIN OPTIONS

GIEMSA STAIN

Blood Concentration

BUFFY COAT CONCENTRATION

KNOTT CONCENTRATION

MEMBRANE FILTRATION CONCENTRATION

Algorithm 5.1 Procedure for processing fresh stool for the O&P examination

Algorithm 5.2 Procedure for processing liquid specimens for the O&P examination

Algorithm 5.3 Procedure for processing preserved stool for the O&P examination—two‐vial collection kit

Algorithm 5.4 Procedure for processing SAF‐preserved stool for the O&P examination

Algorithm 5.5 Procedure for the use of Total‐Fix (universal fixative, single‐vial system) (this fixative contains no mercury, no PVA, and no formalin)*

Algorithm 5.6 Use of various fixatives and their recommended stains

Algorithm 5.7 Ordering algorithm for laboratory examination for intestinal parasites

Table 5.1 Body sites, specimens, and recommended stains

Table 5.2 Approaches to stool parasitology: test ordering

Table 5.3 Laboratory test reports: notes and optional comments

Table 5.4 Parasitemia determined from conventional light microscopy: clinical correlation

SECTION 6: Commonly Asked Questions about Diagnostic Parasitology

Stool Parasitology

Specimen Collection

Specimen Processing

Diagnostic Methods

Stool Immunoassay Options

MOLECULAR TEST PANELS (FDA CLEARED)

A.APTIMA Trichomonas vaginalis Assay

B. Affirm VPIII Microbial Identification Test

C. Cepheid Xpert TV Assay for Trichomonas vaginalis from Men and Women

D. BD MAX Enteric Parasite Panel

E. BioFire FilmArray Gastrointestinal Panel

F. Luminex (Verigene II GI Flex Assay; Includes Parasites)

G. Other Pending Molecular Tests

Organism Identification

Reporting

Proficiency Testing

Tissues or Fluids

Blood

Specimen Collection

Specimen Processing

Diagnostic Methods

Organism Identification

Reporting

Proficiency Testing

General Questions

SECTION 7: Parasite Identification

PROTOZOA  •  Amebae (Intestinal)

Entamoeba histolytica

Entamoeba histolytica/Entamoeba dispar

Comments on Entamoeba moshkovskii and Entamoeba bangladeshi

Entamoeba bangladeshi

Entamoeba hartmanni

Entamoeba coli

Entamoeba gingivalis

Entamoeba polecki

Endolimax nana

Iodamoeba bütschlii

Blastocystis spp. (formerly Blastocystis hominis)

PROTOZOA • Flagellates (Intestinal)

Giardia lamblia (G. duodenalis, G. intestinalis)

Dientamoeba fragilis

Chilomastix mesnili

Pentatrichomonas hominis

Enteromonas hominis

PROTOZOA • Ciliates (Intestinal)

Balantidium coli

PROTOZOA • Apicomplexa (Intestinal)

Cryptosporidium spp.

PROTOZOA • Coccidia (Intestinal)

Cyclospora cayetanensis

Cystoisospora (formerly Isospora) belli

PROTOZOA • Microsporidia (Intestinal)

Enterocytozoon bieneusi

Encephalitozoon intestinalis

PROTOZOA • Sporozoa (Blood and Tissue)

Plasmodium vivax

Plasmodium falciparum

Plasmodium malariae

Plasmodium ovale wallickeri Plasmodium ovale curtisi

Plasmodium knowlesi

Malaria

Babesia spp. (Babesia microti, B. duncani, B. divergens, B. venatorum)

Toxoplasma gondii

PROTOZOA • Flagellates (Blood and Tissue)

Leishmania spp.

Trypanosoma brucei gambiense (West)

Trypanosoma cruzi

PROTOZOA • Amebae (Other Body Sites)

Naegleria fowleri

Acanthamoeba spp.

PROTOZOA • Flagellates (Other Body Sites)

Trichomonas vaginalis

NEMATODES • Intestinal

Ascaris lumbricoides

Trichuris trichiura Capillaria philippinensis

Necator americanus

Strongyloides stercoralis

Enterobius vermicularis

NEMATODES • Tissue

Ancylostoma braziliense

Toxocara canis

Dracunculus medinensis

Trichinella spiralis

NEMATODES • Blood and Tissue

Filarial Worms

CESTODES • Intestinal

Taenia saginata

Taenia solium

Diphyllobothrium latum

Hymenolepis (Rodentolepis) nana

Hymenolepis diminuta

Dipylidium caninum

CESTODES • Tissue

Echinococcus granulosus

TREMATODES • Intestinal

Fasciolopsis buski

TREMATODES • Liver and Lungs

Paragonimus westermani

Fasciola hepatica

Clonorchis (Opisthorchis) sinensis (Chinese liver fluke)

TREMATODES • Blood

Schistosoma spp. (Schistosoma mansoni, S. haematobium, S. japonicum, S. mekongi, S. malayensis, S. intercalatum)

SECTION 8: Common Problems in Parasite Identification

SECTION 9: Identification Aids

Abbreviations

DIAGNOSTIC CONSIDERATIONS

PROTOZOA

HELMINTHS

BLOOD PARASITES

Index

End User License Agreement

List of Tables

Chapter 1

Table 1.1 Common features of the neglected tropical diseases

a

Chapter 2

Table 2.1 Classification of human parasites

Table 2.2 Cosmopolitan distribution of common parasitic infections

a

Table 2.3 Body sites and possible parasites recovered

a

Chapter 3

Table 3.1 Fecal specimens for parasites: options for collection and processin...

Table 3.2 Approaches to stool parasitology: test ordering

Table 3.3 Preservatives and procedures commonly used in diagnostic parasitolo...

Table 3.4 Advantages of thin and thick blood films

a

Table 3.5 Advantages and disadvantages of buffy coat films

Table 3.6 Potential problems of using EDTA anticoagulant for the preparation ...

Table 3.7 Body sites and possible parasites recovered (trophozoites, cysts, o...

Chapter 4

Table 4.1 Body sites, procedures and specimens, recommended methods and relev...

Table 4.2 Serologic, antigen, and probe tests used in the diagnosis of parasi...

Chapter 5

Table 5.1 Body sites, specimens, and recommended stains

Table 5.2 Approaches to stool parasitology: test ordering

Table 5.3 Laboratory test reports: notes and optional comments

Table 5.4 Parasitemia determined from conventional light microscopy: clinical co...

Chapter 6

Table 6.1

Table 6.2

Table 6.3

Chapter 8

Table 8.1 Entamoeba sp. trophozoites versus macrophages

Table 8.2 Entamoeba sp. cysts versus PMNs

Table 8.3 Entamoeba histolytica versus Entamoeba coli precysts and cysts

Table 8.4 Endolimax nana versus Dientamoeba fragilis

Chapter 9

Table 9.1 Rapid diagnostic procedures

Table 9.2 Diagnostic characteristics for organisms in wet mounts (direct or c...

Table 9.3 Diagnostic characteristics for organisms in permanent stained smear...

Table 9.4 Intestinal protozoa: trophozoites of common amebae

Table 9.5 Intestinal protozoa: cysts of common amebae

Table 9.6 Intestinal protozoa: trophozoites of less common amebae

Table 9.7 Intestinal protozoa: cysts of less common amebae

Table 9.8 Morphologic criteria used to identifyBlastocystis spp.

Table 9.9 Intestinal protozoa: trophozoites of flagellates

Table 9.10 Intestinal protozoa: cysts of flagellates

Table 9.11 Intestinal protozoa: ciliate (Balantidium coli)

Table 9.12 Apicomplexa

Table 9.13 Microsporidia (related to the Fungi): general information

Table 9.14 Microsporidia: recommended diagnostic techniques

Table 9.15 Comparison ofNaegleria fowleri, Acanthamoeba spp., Balamuthia mand...

Table 9.16 Characteristics ofTrichomonas vaginalis

Table 9.17 Key characteristics of intestinal tract and urogenital system prot...

Table 9.18 Normal life spans of the most common intestinal nematodes

Table 9.19 Characteristics of the most common intestinal nematodes

Table 9.20 Tissue nematodes

Table 9.21Trichinella spiralis: life cycle stages and clinical conditions

Table 9.22 Characteristics of human microfilariae

Table 9.23 Characteristics of cestode parasites (intestinal)

Table 9.24 Tissue cestodes

Table 9.25 Characteristics of intestinal trematodes

Table 9.26 Characteristics of liver and lung trematodes

Table 9.27 Human paragonimiasis

Table 9.28 Characteristics of blood trematodes

Table 9.29 Key characteristics of helminths

a

Table 9.30 Malaria characteristics with fresh blood or EDTA‐blood

a

Table 9.31 Potential problems with using EDTA anticoagulant for the preparati...

Table 9.32 Plasmodia in Giemsa‐stained thin blood smears

a

Table 9.33 Relevant issues for handling requests for identification of infect...

Table 9.34 Features of human leishmanial infections

a

Table 9.35 Characteristics of American trypanosomiasis

Table 9.36 Characteristics of East and West African trypanosomiasis

Table 9.37 Key characteristics of blood parasites

a

List of Illustrations

Chapter 1

Figure 1.1 Free‐living amebae on nonnutrient agar seeded with E. coli. Left,...

Figure 1.2 When placed in distilled water (enflagellation test), N. fowleri,...

Chapter 2

Figure 2.1 Representative intestinal amebae (Entamoeba spp.). (Top row) Tro...

Figure 2.2 Intestinal Stramenopiles (Blastocystis spp.). Central‐body forms...

Figure 2.3 Representative intestinal flagellates. Giardia lamblia (also cal...

Figure 2.4 The intestinal ciliate Balantidium coli. Trophozoite (note the l...

Figure 2.5 Apicomplexa (top) and coccidia (bottom). (Top row) Cryptosporidiu...

Figure 2.6 Microsporidia (fungi). Microsporidian spores (note the cross/dia...

Figure 2.7 Protozoa (Naegleria fowleri, top) and amebae (Acanthamoeba spp.,...

Figure 2.8 Flagellates from other body sites. Trichomonas vaginalis (urinar...

Figure 2.9 Blood and tissue Apicomplexa (including Sporozoa): Plasmodium sp...

Figure 2.10 Blood and tissue Apicomplexa (including Sporozoa): Babesia spp....

Figure 2.11 Blood and tissue flagellates: leishmaniae. Examples of cutaneou...

Figure 2.12 Blood and tissue flagellates: trypanosomes. Trypanosoma brucei ...

Figure 2.13 Nematodes (intestinal). (Top row) Adult Ascaris lumbricoides ma...

Figure 2.14 Nematodes (tissue). (Row 1) Trichinella sp. encysted larva in mu...

Figure 2.15 Nematodes (blood and tissue): filarial worms. (Top row) Wucherer...

Figure 2.16 Cestodes (intestinal). (Row 1) Taenia sp. egg (without special s...

Figure 2.17 Cestodes (tissue). (Top row) Brain with multiple cysticerci of

Figure 2.18 Trematodes (intestinal). Fasciolopsis buski adult fluke (giant ...

Figure 2.19 Trematodes (liver and lungs). Clonorchis sinensis adult fluke (...

Figure 2.20 Trematodes (blood). Adult schistosomes (note that the slender f...

Figure 2.21 Pentastomids (tongue worms). (Left) Male (small) and female (la...

Figure 2.22 Acanthocephala (thorny‐headed worms). (Left) Macracanthorhynchu...

Chapter 4

Figure 4.1 Direct wet mount or concentration sediment wet mount. Giardia lam...

Figure 4.2 Permanent stained smear (trichrome and iron hematoxylin). (Top ro...

Figure 4.3 Strongyloides stercoralis agar plate culture. Agar plate showing ...

Figure 4.4 Hatched Schistosoma haematobium egg found in urine contaminated w...

Figure 4.5 Enterobius vermicularis (pinworm) eggs collected using cellophane...

Figure 4.6 Entero‐Test capsule for sampling duodenal contents. Left to right...

Figure 4.7 Nuclepore membrane filtration for recovery of microfilariae. Filt...

Figure 4.8 Xenodiagnosis.

Figure 4.9 Intradermal test showing a positive result.

Chapter 5

Figure 5.1 Ocular micrometer (top scale) and stage micrometer (bottom scale)...

Figure 5.2 Method of scanning a direct wet film preparation with the 10× obj...

Figure 5.3 Direct wet mount and/or concentration wet mount organism images (...

Figure 5.4 Fecal concentration procedures. Various layers are seen in tubes ...

Figure 5.5 Method used to remove surface film in the zinc sulfate flotation ...

Figure 5.6 Intestinal protozoa stained with Wheatley's trichrome stain. (Top...

Figure 5.7 Intestinal protozoa stained with iron hematoxylin stain. Top row,...

Figure 5.8 Iron hematoxylin stain incorporating the carbol fuchsin step. Not...

Figure 5.9 Modified acid‐fast stains. (Top row) Cryptosporidium sp. oocysts;...

Figure 5.10 Rapid hot safranin stain of Cyclospora cayetanensis oocysts.

Figure 5.11 Auramine O staining for Apicomplexa. (Top row) Cryptosporidium s...

Figure 5.12 Modified trichrome stain for microsporidian spores. Weber green ...

Figure 5.13 Fecal immunoassay formats. EIA plate for Giardia with positive y...

Figure 5.14 Culture methods for the recovery of larval‐stage nematodes, incl...

Figure 5.15 Baermann apparatus. (Illustration by Nobuko Kitamura; reprinted ...

Figure 5.16 Agar culture method for Strongyloides stercoralis. (1) Agar plat...

Figure 5.17 Enterobius vermicularis (pinworm). E. vermicularis eggs; E. verm...

Figure 5.18 Enterobius vermicularis eggs on cellophane tape. Note football‐s...

Figure 5.19 Diagram of a commercial kit (Evergreen Scientific) for use in sa...

Figure 5.20 Nuclepore filtration system for various human parasites. (Left) ...

Figure 5.21 Method of thin blood film preparation. (a) Position of spreader ...

Figure 5.22 Method of thick‐thin combination blood film preparation. (a) Pos...

Figure 5.23 Buffy coat stained blood films. Leishmania donovani (note the sm...

Figure 5.24 Stained examples of Plasmodium spp. parasites. Plasmodium vivax,...

Algorithm 5.1 Procedure for processing fresh stool for the O&P examination...

Algorithm 5.2 Procedure for processing liquid specimens for the O&P examinat...

Algorithm 5.3 Procedure for processing preserved stool for the O&P examinati...

Algorithm 5.4 Procedure for processing SAF‐preserved stool for the O&P exami...

Algorithm 5.5 Procedure for the use of Total‐Fix (universal fixative, single...

Algorithm 5.6 Use of various fixatives and their recommended stains

Algorithm 5.7 Ordering algorithm for laboratory examination for intestinal p...

Algorithm 5.8 Procedure for processing blood specimens for examination

Chapter 6

Figure 6.1 Traditional method for preparing a thin blood film. The blood can...

Figure 6.2 Poorly prepared thin and thick blood films (dirty slides, oil on ...

Figure 6.3 Method of thick‐thin combination blood film preparation. (a) Posi...

Figure 6.4 Babesia spp. The ring configuration called the Maltese cross (cir...

Figure 6.5 General diagram of a rapid malaria test. (Top) The negative test ...

Chapter 7

Figure 7.1 Entamoeba histolytica (true pathogen). (Top row) The first two dr...

Figure 7.2 Entamoeba histolytica/E. dispar. (Top row) Entamoeba histolytica/E...

Figure 7.3 Entamoeba hartmanni. (Top row) Entamoeba hartmanni trophozoite (no...

Figure 7.4 Entamoeba coli. (Row 1) Entamoeba coli trophozoite (no ingested RB...

Figure 7.5 Entamoeba gingivalis (trophozoites only) and Entamoeba polecki. (...

Figure 7.6 Endolimax nana. (Top row) Endolimax nana trophozoite (single nucle...

Figure 7.7 Iodamoeba bütschlii. (Top row) Trophozoite (note the cytopla...

Figure 7.8 Blastocystis spp. (Top row) Drawing of the Blastocystis central‐b...

Plate 7.A (Row 1) Entamoeba histolytica trophozoite; Entamoeba histolytica/E....

Plate 7.B (Row 1) Blastocystis central‐body forms (note the peripheral nucle...

Figure 7.9 Giardia lamblia. (Top row) Two Giardia trophozoites (note the two...

Figure 7.10 Dientamoeba fragilis. Drawings: Forms of Dientamoeba fragilis: c...

Figure 7.11 Chilomastix mesnili. (Left) Drawings of a Chilomastix mesnili tr...

Figure 7.12 Pentatrichomonas hominis, Trichomonas tenax . Photograph of P. ho...

Figure 7.13 Enteromonas hominis, Retortamonas intestinalis. (Top row) Drawin...

Figure 7.14 Balantidium coli. (Top row) Drawings of Balantidium coli trophoz...

Plate 7.C (Row 1) Giardia lamblia trophozoites. (Row 2) G. lamblia trophozoite...

Plate 7.D (Row 1) Dientamoeba fragilis trophozoites. (Row 2) D. fragilis troph...

Figure 7.15 Cryptosporidium spp. (Top row) Artist's rendering of Cryptospori...

Figure 7.16 Cyclospora cayetanensis. (Row 1) (a) Unsporulated oocyst with un...

Figure 7.17 Cystoisospora belli. (Top row) Drawing of an immature oocyst; dr...

Figure 7.18 Enterocytozoon bieneusi. (Left) Drawing of an infective spore (1...

Figure 7.19 Encephalitozoon spp. (Left) Drawing of an infective spore (1.5 t...

Plate 7.E (Row 1) Cryptosporidium oocysts in a spinach leaf stoma (SEM, cour...

Plate 7.F (Row 1) Cystoisospora belli oocysts (from left to right: modified ...

Figure 7.20 Method for preparation of thin blood film. (A) Position of sprea...

Figure 7.21 Plasmodium vivax. (Row 1) Illustrations of early ring form, deve...

Figure 7.22 Plasmodium falciparum. (Row 1) Illustrations of ring forms. (Row...

Figure 7.23 Plasmodium malariae. (Row 1) Illustrations of young rings (first...

Figure 7.24 Plasmodium ovale. (Row 1) Illustrations of young and developing ...

Figure 7.25 Plasmodium knowlesi. (Top row) Ring form; developing trophozoite...

Figure 7.26 Babesia spp. (Top row) Babesia rings (note the Maltese cross form...

Figure 7.27 Toxoplasma gondii. (Top row) T. gondii tachyzoites in bone marrow...

Plate 7.G Plasmodium vivax. (Row 1) Developing ring forms (in some RBCs ther...

Plate 7.H Plasmodium falciparum. (Row 1) Developing ring forms of P. falcipa...

Plate 7.I Plasmodium malariae. (Rows 1 and 2) Developing trophozoites (note ...

Plate 7.J Plasmodium ovale . (Rows 1 and 2) Developing trophozoites. The para...

Plate 7.K Plasmodium knowlesi . (Row 1) P. knowlesi ring forms (the third imag...

Plate 7.L Babesia spp. (Rows 1, 2, and 3) Examples of blood films containing...

Figure 7.28 Leishmania spp. (Left) Drawing of amastigotes in bone marrow (vi...

Figure 7.29 African trypanosomes (Trypanosoma brucei gambiense, T. brucei rh...

Figure 7.30 Trypanosoma cruzi. T. cruzi amastigotes in cardiac tissue and ty...

Plate 7.M (Row 1) Leishmania amastigotes within macrophage (note the nucleus ...

Plate 7.N (Row 1) Trypanosoma brucei gambiense or T. brucei rhodesiense (Afri...

Figure 7.31 Naegleria fowleri. (Top row) Primary amebic meningoencephalitis ...

Figure 7.32 Free‐living amebae. (Top row) Acanthamoeba trophozoite (not...

Figure 7.33 Trichomonas vaginalis. Drawing of T. vaginalis (note that the un...

Figure 7.34 Ascaris lumbricoides. (Top row) Illustrations of unfertilized eg...

Figure 7.35 Trichuris trichiura. (Top row) Drawings of T. trichiura eggs; ad...

Figure 7.36 Hookworm. (Top row) Drawing of a hookworm egg (Necator americanu...

Figure 7.37 Strongyloides stercoralis. (Top row) Short mouth opening of Stro...

Figure 7.38 Enterobius vermicularis. (Top row) Adult male and female pinworm...

Figure 7.39 Cutaneous larva migrans. (Top row) Linear tracks on the top of t...

Figure 7.40 Toxocara spp. (Top row) Toxocara eggs (note the dimpled shells [s...

Figure 7.41 Dracunculus medinensis. (Top) Dracunculus medinensis (Guinea wor...

Figure 7.42 Trichinella spp. (Top row) Drawing of an encysted larva (illustr...

Figure 7.43 Filarial infections. Diagrams of human microfilariae—Wuchereria ...

Plate 7.O (Row 1) Brugia malayi microfilaria (note the pink sheath and two te...

Plate 7.P (Row 1) Onchocerca volvulus (nodule on scalp); O. volvulus (organis...

Figure 7.44 Taenia saginata. Drawings depicting the scolex, egg, and gravid ...

Figure 7.45 Taenia solium. (Top row) Drawings depicting the scolex, egg, and...

Figure 7.46 Racemose cysticercosis . Racemose form of Taenia solium cysticerc...

Figure 7.47 Diphyllobothrium latum. (Top row) Drawings depicting the scolex,...

Figure 7.48 Hymenolepis nana. (Top row) Drawing depicting the H. nana egg; p...

Figure 7.49 Hymenolepis diminuta. (Top row) Drawing depicting the H. diminut...

Figure 7.50 Dipylidium caninum. (Row 1) Drawings depicting the scolex, gravi...

Figure 7.51 Echinococcus granulosus. (Top row) Adult worm (three proglottids...

Plate 7.Q (Row 1) Taenia saginata scolex; T. saginata gravid proglottid; Taen...

Plate 7.R (Top row) Echinococcus granulosus hydatid cyst (cystic disease) con...

Figure 7.52 Fasciolopsis buski. F. buski adult worm; F. buski egg (the opercu...

Figure 7.53 Paragonimus spp. (Top row) Drawings showing Diphyllobothrium lat...

Figure 7.54 Fasciola hepatica. (Top row) Drawing of F. hepatica egg (the ope...

Figure 7.55 Clonorchis sinensis. (Top row) Drawing and photographs of eggs o...

Figure 7.56 Schistosoma spp. (Row 1) Scanning electron micrograph of male an...

Chapter 8

Figure 8.1 (a) Entamoeba histolytica/E. dispar trophozoite. Note the evenly ...

Figure 8.2 (a) Entamoeba histolytica/E. dispar trophozoite. Note the evenly ...

Figure 8.3 (a) Entamoeba histolytica/E. dispar trophozoite. Again, note the ...

Figure 8.4 (Top row) Entamoeba histolytica/E. dispar trophozoite (wet mount; n...

Figure 8.5 (a) Entamoeba histolytica trophozoite. Note the evenly arranged n...

Figure 8.6 (Top row) Entamoeba histolytica trophozoites on permanent stained s...

Figure 8.7 (a) Entamoeba histolytica/E. dispar precyst. Note the enlarged nu...

Figure 8.8 (a) Entamoeba histolytica/E. dispar mature cyst. Note that the fo...

Figure 8.9 (Top row) PMNs. Note the appearance of “multiple nuclei” in the ce...

Figure 8.10 (Top row) Entamoeba coli precyst with two enlarged nuclei, one on ...

Figure 8.11 (a) Endolimax nana trophozoite. This organism is characterized b...

Figure 8.12 (a) Endolimax nana trophozoite. Note that the karyosome is large...

Figure 8.13 Endolimax nana trophozoites. Note the tremendous nuclear variatio...

Figure 8.14 Dientamoeba fragilis trophozoites. (Rows 1 to 3) Some organisms h...

Figure 8.15 (a) Endolimax nana trophozoite. Note the large karyosome surroun...

Figure 8.16 (Top row) Endolimax nana trophozoite (note the large single karyos...

Figure 8.17 (Top row) (a) RBCs on a stained fecal smear. Note that the cells ...

Figure 8.18 (Top row) (a) Entamoeba histolytica/E. dispar cyst. Note the shri...

Figure 8.19 Various structures that may be seen in stool preparations. (Row ...

Figure 8.20 Giardia lamblia and Cystoisospora belli. Two images of Giardia la...

Figure 8.21 (Rows 1 and 2) Stain deposition on the surface of uninfected RBCs...

Figure 8.22 Male Plasmodium gametocyte (microgametocyte) undergoing exflagel...

Figure 8.23 (Left) Histoplasma capsulatum. (Middle) Leishmania donovani. Note t...

Figure 8.24 Bronchial epithelium cells. When these cells disintegrate, the c...

Figure 8.25 (a) Plasmodium falciparum rings. Note the two rings in the RBC. ...

Figure 8.26 (Top row) Plasmodium falciparum rings (note the “clean” morphology...

Figure 8.27 (Top) Root hair. (Middle) Root hair. Note that there is no intern...

Figure 8.28 Various artifacts that may be seen in stool preparations (wet mo...

Figure 8.29 Various artifacts that may be seen in stool preparations (wet mo...

Figure 8.30 Various types of pollen grains and a root hair (fifth row, left)...

Figure 8.31 White blood cells in a stained blood film. (1) Lymphocytes. (2) ...

Figure 8.32 (Top row) Charcot‐Leyden (CL) crystals in trichrome‐stained fecal...

Chapter 9

Figure 9.1 Intestinal amebae of humans. (Top row) Trophozoites. Entamoeba hi...

Figure 9.2 Intestinal and urogenital flagellates of humans. (Top row) Tropho...

Figure 9.3 Helminth eggs depicted in the order of size (smallest to largest)...

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PRACTICAL GUIDE TODiagnosticParasitology

THIRD EDITION

Lynne S. Garcia, MS, MT(ASCP), CLS(NCA), F(AAM)

LSG & Associates, Santa Monica, California

Copyright © 2021 American Society for Microbiology. All rights reserved. Copublication by the American Society for Microbiology and John Wiley & Sons, Inc. No part of this publication may be reproduced, stored in a retrieval system, or transmitted in any form or by any means electronic, mechanical, photocopying, recording, scanning, or otherwise, except as permitted by law. Advice on how to reuse material from this title is available at http://wiley.com/go/permissions.

The right of Lynne Shore Garcia to be identified as the author of this work has been asserted in accordance with law.

Limit of Liability/Disclaimer of Warranty

While the publisher and author have used their best efforts in preparing this book, they make no representations or warranties with respect to the accuracy or completeness of the contents of this book and specifically disclaim any implied warranties or merchantability or fitness for a particular purpose. No warranty may be created or extended by sales representatives or written sales materials. The publisher is not providing legal, medical, or other professional services. Any reference herein to any specific commercial products, procedures, or services by trade name, trademark, manufacturer, or otherwise does not constitute or imply endorsement, recommendation, or favored status by the American Society for Microbiology (ASM). The views and opinions of the author(s) expressed in this publication do not necessarily state or reflect those of ASM, and they shall not be used to advertise or endorse any product.

Editorial Correspondence: ASM Press, 1752 N Street, NW, Washington, DC 20036-2904, USA Registered Offices: John Wiley & Sons, Inc., 111 River Street, Hoboken, NJ 07030, USA For details of our global editorial offices, customer services, and more information about Wiley products, visit us at www.wiley.com.

Wiley also publishes its books in a variety of electronic formats and by print-on-demand. Some content that appears in standard print versions of this book may not be available in other formats.

Library of Congress Cataloging‐in‐Publication data

Names: Garcia, Lynne Shore, author. | American Society for Microbiology.

Title: Practical guide to diagnostic parasitology / Lynne S. Garcia.

Description: 3rd edition. | Hoboken, NJ : Wiley, 2021. | Includes bibliographical references and index.

Identifiers: LCCN 2021001642 (print) | LCCN 2021001643 (ebook) | ISBN 9781683670391 (hardback) | ISBN 9781683670407 (adobe pdf) | ISBN 9781683673620 (epub)

Subjects: MESH: Parasitic Diseases‐diagnosis | Clinical Laboratory Techniques

Classification: LCC QR255 (print) | LCC QR255 (ebook) | NLM WC 695 | DDC 616.9/6075-dc23

LC record available at https://lccn.loc.gov/2021001642

LC ebook record available at https://lccn.loc.gov/2021001643

Cover image: A group of red blood cells (magenta) with the central cell infected by Plasmodium falciparum, the parasite that causes the deadliest form of malaria. Courtesy of Dr. Michał Pasternak, WEHI (Walter and Eliza Hall Institute), https://imaging.wehi.edu.au/our‐work/new‐views‐malaria‐parasites.

Cover and interior design: Susan Brown Schmidler

I dedicate the third edition of this practical guide to Robyn Shimizu. Robyn and I have worked together for many years, performing bench work; training students, post‐doctoral fellows, and medical residents; presenting workshops and seminars; handling consult; performing studies; and preparing manuscripts for publication. It's been a wonderful collaboration and sharing of information throughout our careers. A very special thanks to Robyn for sharing this educational adventure; hopefully our many students have found these contributions helpful.

Preface

As we move forward into the 21st century, the field of diagnostic medical parasitology continues to see some dramatic changes, including newly recognized pathogens, changing endemicity of familiar pathogens, disease control challenges, geographic and climate changes that support the spread of parasitic disease, new methodologies, expansion of diagnostic testing, and an ongoing review of the approach to and clinical relevance of this type of diagnostic testing on patient care within the managed care environment, as well as the world as a whole.

The third edition of the Practical Guide to Diagnostic Parasitology is organized to provide maximum help to the user, particularly from the bench use perspective. New aspects of the field have been addressed throughout, and many new figures and plates have been added, including extensive color images. All of the changes for this edition are based on the need for readers to update their information related to diagnostic medical parasitology and specifically issues involving laboratory techniques. With continued emphasis on regulatory requirements related to chemical disposal and the use of mercury compounds, laboratories are being required to develop skills using substitute fixatives that are prepared without the use of mercury‐based compounds. In most cases, organism identification is comparable; an example of a rare exception is one in which the number of organisms present is quite low. This is a prime example of a change where “different” has been acceptable and relevant, not necessarily “good” or “bad.” It is also important to remember the large number of variables relevant to diagnostic parasitology testing.

Section 1 of this new edition, on the philosophy and approach to diagnostic parasitology, has been expanded to include discussions on neglected tropical diseases, the impact of global climate change, population movements, potential outbreak testing, the development of laboratory test menus, and the risk management issues related to “stat” testing. The Section 2 discussion of organism classification and relevant tables has been expanded and updated to provide the user with current information related to changes in nomenclature and the overall importance of the various parasite categories to human infection.

In Section 3, expanded information on stool specimen fixatives and testing options has been provided. This information is valuable for any laboratory that is reviewing collection and testing options related to fixative compatibility with the routine ova and parasite examination, as well as fecal immunoassays and the newer molecular parasite panels. It is always important to check the literature from the relevant company to confirm the FDA status of any new product. The discussion on blood collection, including the pros and cons of current changes from finger‐stick blood to venipuncture, has been greatly enhanced, particularly related to potential problems with blood parasite morphology and lag time issues. Additional tables serve to summarize much of this new information.

New tables and information have been added to Sections 4 and 5, including a number of new algorithms. Section 6 is one of the most important sections in the book, with extensive revisions related to the most commonly asked questions regarding diagnostic parasitology methods. Additional techniques have been included (molecular test panels), as well as new information related to reporting results and the importance of report comments. This section’s “FAQ” format makes it easy for the reader to use the expanded information.

Section 7 has been greatly expanded, including the addition of extensive color figures. Figures and life cycle diagrams have also been expanded and updated. Section 8 presents information on potential problems with organism differentiation from one another and from possible artifacts. The section has been significantly expanded with new tables and image plates to illustrate these differences. Section 9 contains numerous tables and a set of identification keys that summarize identification aids and organism characteristics. As with earlier sections, the information is presented to assist the bench microbiologist with routine diagnostic testing methods.

Many laboratories are reviewing all microbiological services, and specific questions are being asked related to diagnostic parasitology options. Some of these questions include the following: what laboratories should be performing this type of testing, when should testing be performed, what tests should be performed, and what factors should be considered when developing test menus. There are also ongoing discussions related to the development of automated parasite panels and how the use of these panels could impact many of the routine procedures now in use.

Laboratories are also reviewing specimen collection options, particularly as they relate to their geographic area and types of patients serviced. This kind of analysis is beneficial to all concerned, not only in helping laboratories to understand the specimen collection options, but how they relate to test orders, diagnostic testing, and results impacting patient care.

With changes in collection, testing, reporting, and interpretation options, it is critical to remember that this information needs to be shared with the laboratory's client base, particularly if the test orders and results are to be used for the best‐quality patient care. Although there are many ways to approach diagnostic parasitology testing, it is mandatory that the laboratory and user both understand the pros and cons of the methods selected and the importance of test menu names, particularly in terms of procedure limitations and test name relevance for billing functions. The use of different approaches to parasitology diagnostic testing is acceptable; however, the benefits and drawbacks must be thoroughly understood by all participants. There may be legitimate reasons why different approaches are used by different laboratories; however, cost containment must not be the sole factor in selecting methods.

Another consideration is the fact that not all clinical laboratories will continue to perform diagnostic parasitology testing. This may be due to financial considerations, lack of skilled personnel, etc. With increased emphasis on cross‐trained individuals, the technical expertise required to identify these parasites by using routine microscopy may be lacking. Even with the use of molecular diagnostics, these tests are not capable of covering the entire spectrum of organisms that may be present as pathogens. However, as more of these newer automated molecular panels become commercially available, the use of nonmicroscopic methods will increase. Many laboratories now include both the ova and parasite examination and various fecal immunoassays on their routine test menus; on the basis of patient histories and symptoms, appropriate orders may focus on one or the other of these options. An important consideration in deciding whether to send out parasitology testing, or to maintain the testing in‐house, relates to stat testing (collection, processing, testing, reporting of thick and thin blood films, and the examination of cerebrospinal fluid and other specimens for the presence of free‐living amebae). These tests must be handled as stat; the time required from collection to reporting must be considered prior to moving these procedures off‐site or sending specimens to a reference laboratory.

Based on the many changes in clinical laboratories within the past few years and many years' experience with teaching and diagnostic bench work, it is my hope that the information contained in this third edition will provide valuable information for the user. This guide is not designed to serve as a diagnostic parasitology text or to contain all possible diagnostic test options, but to help the user make some sense of a field for which training has become almost nonexistent. I have included a section on commonly asked questions about diagnostic medical parasitology and hope that this discussion will be of practical value to the user; the answers to some of these questions are often difficult to find, even in a more comprehensive book. Again, let me emphasize that different approaches to laboratory work are not always “good” or “bad.” The key to success is making sure that both the laboratory and clients understand the pros and cons of each collection, testing, and reporting option and that educational information is provided for all clients on an ongoing basis. Two of the most important functions of the clinical laboratory in the future will be educational and consultative, particularly when laboratory services are within the microbiology area. The importance of well‐trained bench microbiologists cannot be underestimated.

A final point is that infectious diseases, particularly parasitic infections, play a huge role in the world's overall health and economy. As travel increases, we anticipate seeing many more people who will be infected with parasites that may not be endemic to the specific area where they live. Continued vector and disease control efforts will remain on the high‐priority list, especially when seen within the context of global health. It is hoped that parasitologists and microbiologists, including those who have diagnostic skills, will be available to support these global initiatives.

ACKNOWLEDGMENTS

I thank the hundreds of colleagues over the past years who have shared their thoughts and suggestions regarding this fascinating field of diagnostic parasitology. There are too many of you to name—you all know who you are, and we all recognize the pitfalls, as well as the fun, in providing diagnostic services in this field of microbiology.

I also thank the many bench techs and microbiologists who have tackled this field over the last 40+ years, including those who attended workshops and seminars; your contributions to the growth and expansion of diagnostic parasitology have been significant. Discussions of questions asked, problems for resolution, and reviews of testing options have been invaluable in shaping our approach to diagnostics in this field. This type of interaction helped all of us keep an open mind when reviewing options and possible new ways to approach the work.

Over the years, our association with many companies has also been extremely valuable in helping to understand test development, test trials, and relevance of results to the ultimate user within the diagnostic laboratory. Again, these interactions have helped maintain some balance and perspective on new options and their relevance to improved patient care.

A challenge to all of us who are still actively working in this area of diagnostic microbiology: Serve as a mentor to some of the young people entering the field of microbiology. The number of personnel trained in this field continues to decline. Until parasite morphology is no longer required for differentiation and diagnosis, skilled microscopists will remain valuable members of the microbiology team and mandatory for the practice of diagnostic medical parasitology.

I would also like to thank Christine Charlip, the Director of ASM Press (now partnered with John Wiley & Sons to copublish ASM Press titles), and members of the editorial staff, including the developmental editor, Ellie Tupper, the copyeditor, Jennifer Schaffer, and Editorial Rights Coordinator Lindsay Williams; they are outstanding professionals. Their many contributions always help the author “look good,” and I appreciate their collaboration. Perhaps retirement will just have to wait a while longer.

Above all, my very special thanks go to my husband, John, for his love and support for this and other projects over the years. I could never have taken on these challenges without his help, understanding, and wonderful sense of humor.

Note: Images in this book credited to CDC PHIL and CDC DpDx were obtained by courtesy of the Centers for Disease Control and Prevention Public Health Image Laboratory (https://phil.cdc.gov/) and CDC DpDx – Laboratory Identification of Parasites of Public Health Concern (https://www.cdc.gov/dpdx/index.html).

About the Author

Lynne Shore Garcia is the director of LSG & Associates, a firm providing training, teaching, and consultation services for diagnostic medical parasitology and health care administration. A former manager of the UCLA Clinical Microbiology Laboratory, she is a sought‐after speaker (nationally and internationally) and author of hundreds of articles, book chapters, and books including two ASM Press books, Clinical Laboratory Management, Second Edition and Diagnostic Medical Parasitology, Sixth Edition. She served as Editor‐in‐Chief of the ASM Press Clinical Microbiology Procedures Handbook, Third Edition, and was a senior editor for ASM's Clinical Microbiology Reviews journal. She serves as a reviewer for a number of journals, provides consulting to the CAP Microbiology Resource Committee, was chair of the CLSI Parasitology Subcommittee, and is a Fellow of The American Academy of Microbiology. Lynne is the 2009 recipient of the ASM bioMérieux Sonnenwirth Award for Leadership in Clinical Microbiology.

SECTION 1Philosophy and Approach to Diagnostic Parasitology

Neglected Tropical Diseases

The term “neglected tropical diseases” (NTDs) was first used in the early 2000s, primarily reflecting the lack of research funds and limited interest of the health care and pharmaceutical industries in investing in affordable drugs for these diseases. One of the earliest conferences on NTDs was organized by Médicins sans Frontières in early 2002 (1). From 2003 to 2007, key steps were taken to develop a framework for tackling NTDs in a coordinated and integrated way (2). In 2005 and 2006, two important articles (3, 4) provided a list of 15 NTDs, 13 of which were deemed of particular importance in terms of annual mortality rates and global burden. This list of 15 diseases formed the initial scope of PLoS Neglected Tropical Diseases. Included were nine helminth infections (cysticercosis/taeniasis, drancunculiasis [guinea worm], echinococcosis [added by WHO], foodborne trematodiasis, lymphatic filariasis, onchocerciasis, schistosomiasis, the three main soil‐transmitted helminthiases [ascariasis, hookworm infection, and trichuriasis]), three protozoal infections (Chagas’ disease, human African trypanosomiasis, and leishmaniasis), scabies and other ectoparasite infections (added by WHO), and three bacterial infections (Buruli ulcer, leprosy, and trachoma) (Table 1.1) (2).

A subsequent review by Hotez and colleagues titled “Control of Neglected Tropical Diseases,” published in the New England Journal of Medicine in 2007, clearly demonstrated that the term “neglected tropical diseases” had become mainstream (5). In October 2007, the Public Library of Sciences published the inaugural issue of a new open‐access journal, PLoS Neglected Tropical Diseases(6). As of mid‐November 2020, more than 8,200 original research papers, editorials, expert opinions, viewpoints, and other magazine‐type articles on NTDs have been published.

In a publication from 2017, Dr. Hotez discussed the tremendous progress towards neglected tropical disease control or even elimination. However, there are important gaps, nine of which are discussed below; parasitic infections have been emphasized for this list (7).

The first group of problems is linked to the geopolitics of the NTDs.

Regional significance

. There are several NTDs that are very important in the areas where they occur; however, they are generally ignored by the global community. Examples include loiasis in Central Africa and mucocutaneous leishmaniasis in the New World.

Political unrest in the Old World.

Second only to poverty, conflict may have the largest social impact on NTDs. Both cutaneous and visceral leishmaniasis outbreaks are now arising in Syria, Iraq, Afghanistan, Sudan, and South Sudan. Cutaneous leishmaniasis has now reached hyperendemic proportions in current and former ISIS occupation zones, and through forced human emigrations, this NTD may spill over into Lebanon, Turkey, and Jordan.

Political destabilization in the New World.

As Venezuela's health system continues to decline, we have seen the resurgence or reemergence of malaria and NTDs such as Chagas' disease and schistosomiasis.

Climate change and its impact on vector‐borne and zoonotic NTDs.

Climate change, along with poverty, war, and population movements, produces detrimental effects which include the increase and spread of NTDs.

NTDs in “wealthy” nations.

The poor living in the Group of 20 (G20) nations—and also Nigeria (richer than the bottom three or four G20 nations)—account for a majority of the world's disease burden for poverty‐related neglected diseases and NTDs. These numbers include millions of Americans living in the United States with an NTD and significant but often unrecognized levels of poverty and disease in Europe and Australia.

The second group is related to coverage gaps and providing universal access to treatment.

Female genital schistosomiasis.

Female genital schistosomiasis is one of the most common gynecologic conditions of women who live in poverty in Africa and is one of Africa's most important cofactors in its AIDS epidemic.

Patient access to essential drugs for Chagas' disease

. Today, most cases of Chagas' disease occur in Latin America's three large economies: Argentina, Brazil, and Mexico. However, more than 90% of people with

Trypanosoma cruzi

infection do not have access to treatment.

Mass drug administration (preventive chemotherapy).

This includes drugs against scabies, lymphatic filariasis, onchocerciasis, and schistosomiasis.

Research and development (R&D)

for a single approach rather than multiple new approaches for control and disease elimination. For malaria, we will need to pursue several R&D approaches, including new drugs, diagnostics, vaccines, and vector control approaches.

A subsequent article by Dr. Hotez and colleagues in 2018 anticipated a number of challenges related to the emergence and reemergence of these diseases (8). These challenges include stress from climate change and catastrophic weather events, regional conflicts over shifting and limited resources, such as water, and the development and spread of urban helminth infections (schistosomiasis and toxocariasis), foodborne trematode infections, cysticercosis, protozoan infections, and zoonotic toxoplasmosis.

Why Perform Diagnostic Parasitology Testing?

Travel

With the increase in world travel and access to varied populations and geographic areas, we continue to see more “tropical” diseases and infections outside areas of endemicity due to the rapidity with which people and organisms can be transmitted from one place to another. Travel has also become accessible and more affordable for many people throughout the world, including those whose overall health status is in some way compromised. The increased transportation of infectious agents and potential human carriers, particularly via air travel, has been clearly demonstrated during the last few years. It has also been well documented that vectors carrying parasitic organisms can be transported via air travel in baggage and in the unpressurized parts of the plane itself; once released, these infected vectors can then transmit these parasites to humans, even in areas where the infections are not endemic.

Population Movements

In many parts of the world, particularly where conflict is ongoing, there continue to be large population movements. Such movements include refugee migrations to and from areas of endemic parasitic diseases (9, 10). Often, in refugee situations, living conditions are very poor and medical limitations may lead to high levels of parasitic disease and severe illness. Also, migrants may move into countries and geographic areas where serious parasitic infections are generally nonendemic, including Europe and parts of North America. Even if these individuals are uninfected when entering these areas, travel home to visit relatives may result in infections that can be imported when they return.

Control Issues

Control of parasites that cause disease is linked to a number of factors, including geographic location, public health infrastructure, political stability, available funding, social and behavioral customs and beliefs, trained laboratory personnel, health care support teams, environmental constraints, degree of understanding of organism life cycles, and opportunities for control and overall commitment. Often control efforts do not cross political or geographic boundaries; unfortunately, vectors and other carriers of infectious agents do not “play by the rules,” and as a result, these boundaries are meaningless in the context of disease control.

Climate Change

With the continued increase in the global temperature, worldwide climate changes are leading to an overall increase in infectious diseases, vector populations, and ranges of endemicity of both parasites and vectors. Global warming enhances the potential spread of tropical parasitic infections, specifically those due to parasites such as Plasmodium spp., Leishmania spp., and Trypanosoma spp. (11). Examples of vectors whose range is increasing include Anopheles, Aedes, and Culex mosquitoes, hard ticks, and triatomid bugs. Another example is the vectors of schistosomiasis (12).

Epidemiologic Considerations

When newer infectious agents and/or diseases are recognized, there is often very little information available regarding the organism life cycle, potential reservoir hosts, and environmental requirements for survival. Priorities may change, and epidemiologic considerations may have been moved lower on the priority list in areas of the world where they were considered important in the past; unfortunately, funding often plays a role in decisions that impact disease control measures.

Compromised Patients; Potential Sex Bias Regarding Infection Susceptibility; Aging

With the tremendous increase in the number of patients whose immune systems are compromised by underlying illness, chemotherapy, transplantation, AIDS, or age, we are much more likely to see increasing numbers of opportunistic infections, including those caused by parasites. Also, we continue to discover and document organisms that were thought to be nonpathogenic but can cause serious disease in the compromised host. When the possible cause of illness in this patient population is being assessed, the possibility of parasitic infections must be considered as part of the differential diagnosis.

Various studies have revealed a bias toward males regarding susceptibility to and severity of parasitic diseases. Although a number of external factors influence the exposure to infection sources among males and females, one recurrent factor suggests that hormonal influence impacts the simultaneous increase in disease occurrence and hormonal activity during the aging process. However, to date, very few controlled studies have been performed. Hormones are suspected to play a role in parasitic disease processes such as amebiasis, malaria, leishmaniasis, toxoplasmosis, and schistosomiasis (13).

There are various examples of the relationship of parasite pathogenesis and the aging population. Parasite biomass, endothelial activation, and microvascular dysfunction are associated with severe disease in Plasmodium knowlesi malaria and likely contribute to pathogenesis. The association of each of these processes with aging may account for the greater severity of malaria observed in older adults in regions of low endemicity (14).

Approach to Therapy

As new etiologic agents are discovered and the need for new therapeutics increases, more sensitive and specific diagnostic methods to assess the efficacy of newer drugs and alternative therapies will become mandatory. Skilled laboratorians, physicians, public health personnel, and other health care team members will be required to think globally in terms of infectious diseases caused by bacterial, fungal, parasitic, and viral agents, particularly when certain parasitic infections require very specific therapeutic regimens.

Who Should Perform Diagnostic Parasitology Testing?

Laboratory Personnel