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Background Since 1997, human-biting ticks submitted to the Department of Defense Human Tick Test Kit Program (HTTKP) of the US Army Public Health Command have been tested for pathogens by PCR. We noted differences in the phenology and infection prevalence among Ixodes scapularis ticks submitted from military installations in different geographic regions. The aim of this study was to characterize these observed differences, comparing the phenology and pathogen infection rates of I. scapularis submitted from soldiers at two sites in the upper Midwest (Camp Ripley, MN, and Ft. McCoy, WI) and one site in the northeastern US (Ft. Indiantown Gap, PA). Methods From 1997 through 2012, the HTTKP received 1,981 I. scapularis from the three installations and tested them for Anaplasma phagocytophilum, Babesia microti, Borrelia burgdorferi and the Ehrlichiamuris-like (EML) agent using PCR; pathogen presence was confirmed via sequencing or amplification of a second gene target. Pathogen and co-infection prevalence, tick engorgement status, and phenology were compared among installations. Results Greater rates of A. phagocytophilum and Ba. microti infections were detected in ticks submitted from installations in Minnesota than in Wisconsin or Pennsylvania, and the EML agent was only detected in ticks from Minnesota and Wisconsin. Midwestern ticks were also more likely to be co-infected than those from Pennsylvania. Both adult and nymphal ticks showed evidence of feeding on people, although nymphs were more often submitted engorged. Adult I. scapularis were received more frequently in June from Minnesota than from either of the other sites. Minnesota adult and nymphal peaks overlapped in June, and submissions of adults exceeded nymphs in that month. Conclusions There were clear differences in I. scapularis phenology, pathogen prevalence and rates of co-infection among the three military installations. Seasonal and temperature differences between the three sites and length of time a population had been established in each region may contribute to the observed differences. The synchrony of adults and nymphs observed in the upper Midwest has implications for pathogen infection prevalence. The EML agent was only detected in Minnesota and Wisconsin, supporting the previous assertion that this pathogen is currently limited to the upper Midwest.

Summary One critical role played by the pathologist in contemporary breast cancer management is that of providing accurate tumour measurements for staging and prognosis. Numerous factors have the potential to affect the final pathologist-recorded tumour size including diligence in measurement, tumour histology and growth pattern, tissue fixation and processing, prior biopsy and prior therapy. The pathologist should strive to provide three-dimensional tumour measurements in 0.1cm increments from the gross specimen and then confirm these measurements microscopically whenever possible. In the era of breast conservation and emerging ablative therapies, the pathologist should also be familiar with the size-determining capability of various breast imaging techniques and their correlation with pathology. In determining final dimension, the pathologist is in a favourable position to verify or modify the size of a breast tumour based on the sum of information from all pathology specimens and, if appropriate, to correlate size with the patient's imaging studies.

Context.—The correctly completed death certificate provides invaluable personal, epidemiologic, and legal information and should be thorough and accurate. Death certification errors are common and range from minor to severe. Objective.—To determine the frequency and type of errors by nonpathologist physicians at a university-affiliated medical center. Design.—Fifty random patients were identified who died at this academic medical center between January 2002 and December 2003 and did not undergo an autopsy. From medical chart review, clinical summaries were produced. Two pathologists used these summaries to create mock death certificates. The original and mock death certificates were then compared to identify errors in the original certificate. Errors were graded on a I to IV scale, with grade IV being the most severe. Results.—Of the 50 death certificates reviewed, grade I, II, and III errors were noted in 72%, 32%, and 30%, respectively. Seventeen certificates (34%) had grade IV errors (wrong cause or manner of death). Multiple errors were identified in 82% of the death certificates reviewed. Conclusions.—The rate of major (grade IV) death certification errors at this academic setting is high and is consistent with major error rates reported by other academic institutions. We attribute errors to house staff inexperience, fatigue, time constraints, unfamiliarity with the deceased, and perceived lack of importance of the death certificate. To counter these factors, we recommend a multifaceted approach, including an annual course in death certification and discussion of the death certificate for each deceased patient during physician rounds. These measures should result in increased accuracy of this important document.

Goals We present the clinicopathologic features, endoscopic appearance, and ultrastructure of a newly described mesenchymal polyp of the colorectum, termed benign fibroblastic polyp. Study A total of 4 cases from our institution are analyzed using routine histopathology, immunohistochemistry, and electron microscopy, and compared with the original series of 14 cases previously described in the pathology literature. Results Benign fibroblastic polyps appear endoscopically throughout the colorectum as submucosal lesions with a hyperplastic surface component. These lesions are histologically and ultrastructurally distinct, but overlap with other mesenchymal polyps warrants additional immunohistochemical studies for definitive classification. Conclusions Benign fibroblastic polyps of the colorectum comprise a recently described distinct entity of mesenchymal polyps that appear to follow an indolent clinical course. Their etiology is uncertain but may be linked to an exuberant tissue response following mucosal injury. Gastroenterologists should be aware of this new entity that will be increasingly diagnosed by pathologists.

Precise measurement of an invasive breast cancer is crucial for pathological staging and subsequent patient management. Formalin fixation and histological processing may change tissue size, but there is no agreement on which state of the specimen, fresh or fixed, should be used for final tumor measurement. To determine the influence of fixation and processing on breast tumor size, a specific 1-dimensional measurement from 50 invasive breast tumors was recorded in fresh, fixed, and processed/mounted states. Tumors varied in maximum measured dimension from 4 to 20 mm and contained 10% to 90% estimated fibrous tissue (mean, 52.8%). In 96% of cases, there was no difference in measured size between fresh and fixed states. After final processing and mounting, a decrease in size from initial fresh measurement was noted in 40% of cases (mean difference, 2.4 mm; maximum difference, 7 mm). In 9 cases (18%), the measured size increased by a maximum of 3 mm (mean, 1.7 mm) after processing/mounting. Twenty-one cases (42%) showed no change in measurement during the entire fixation and processing protocol. Increases in measured size were attributed largely to tissue expansion during histological sectioning/mounting. One can arguably measure the size of an invasive breast cancer from either the fresh or fixed state without affecting accuracy, but caution should be exercised in relying solely on the microscopic measurements.

We report the finding of Iodamoeba butschlii amebic cysts on a liquid-based anal Pap smear from an HIV-positive male. Iodine staining of the smear confirmed the diagnosis. It is important to distinguish I. butschlii from pathogenic ameobae and other organisms seen on anal Pap smears.

Ehrlichiosis is a tick-borne disease that ranges in severity from asymptomatic infection to fatal sepsis. Ehrlichiosis acquired from transfusion of blood products has not been documented in the literature to date. A case of Ehrlichia ewingii infection likely transmitted by transfusion of leukoreduced platelets is described, and public health implications are discussed.

Tracheobronchial foreign body aspiration is a serious medical problem, with clinical manifestations ranging from acute asphyxiation to insidious lung damage, as demonstrated by the three presented cases. Patient 1 aspirated during dinner, emergency bronchoscopy retrieved pieces of food, and she fully recovered the following day. Patient 2 presented with recurrent pneumonia and a right lower lobe lung abscess. After right lower lobectomy, pathology revealed a foreign object in the right main stem bronchus, a peanut aspirated one year earlier. Patient 3 became unresponsive several days after spinal surgery. The differential diagnosis included myocardial infarction, stroke, and foreign body aspiration. The patient died and necropsy revealed a foreign body in the right main stem bronchus (cooked meat). Thus, foreign body aspiration is not always suspected clinically, and the pathologist may play an important role in making the diagnosis. Histological identification of the aspirated material may be necessary for definitive diagnosis. Therefore, sections of commonly aspirated foods are presented, together with a 10 year history of aspirated objects received by this institution’s surgical pathology department.

Digital imaging is an inexpensive and widely available tool that is used by most pathologists in patient reports, education, publication, diagnosis, and data archival. Its popularity is due, in part, to the ease of modifying, storing, enhancing, and annotating images. Since digital manipulation is essentially undetectable in the final product, it poses the potential risk for fraudulent manipulation and heightens the possibility of unintentional misrepresentation. In an attempt to ensure a high degree of uniformity and quality, and to create a professional standardization amongst pathologists, digital imaging guidelines are proposed for use in general and academic practice.

We report an unusual case of a foreign body removed from the urinary bladder of a 63-year-old male which mimicked a parasitic worm. The foreign body was identified as an artificial fishing worm by morphological comparison to a similar commercially produced product and by infrared spectrum analysis.

This study measured the precision of a quantitative laboratory-developed real-time PCR test for cytomegalovirus performed at three different clinical laboratories that use the same methodology. The overall standard deviation (adjusted for analyte level) was 0.18 log 10 copies/ml, and there was no significant relationship between standard deviation and analytical measuring range.

Herpes simplex virus (HSV) and varicella-zoster virus (VZV) may cause latent infection of the same peripheral nerve ganglia. However, there are no large studies addressing the frequency of concurrent HSV/VZV PCR positivity from the same anatomic location. In an eight-year retrospective study, we observed 1.3% dual positivity from dermal, genital, and oral mucosal sources.

Six cases of amoebic encephalitis admitted to the National Institute of Neoplastic Diseases between the years 1994-2010 in Peru are reported. These cases were admitted for clinical suspicion of malignant primary brain tumor and one orbital-nasal sarcoma. All cases came from coastal regions; three were less than 24 years of age and four were male. The most common symptoms were headache and seizures. Three cases had more than one brain lesion. Stereotactic biopsy was performed in three patients and the differential pathological diagnosis in two cases was glioma of high and low grade. It was possible to confirm the diagnosis using molecular techniques in paraffin-embedded samples in three cases. All patients died within 15 days of admission to the institution. Amoebic encephalitis may be erroneously interpreted as a cerebral neoplasm, causing delay in the management of the infection.

Background The World Health Organization has identified 17 neglected tropical diseases (NTDs) that disproportionately affect the world's poorest populations. The neurologic aspects of many of these NTDs have received relatively little attention. Methods A review was performed in PubMed (MedLine) for each NTD by disease name, name of its causative organism, and neurology, neurosurgery, neurologist, brain, spinal cord, peripheral nerve, muscle, nervous system, encephalitis, meningitis, encephalopathy, stroke, neuropathy, and myopathy (1968-Sept. 2013). The Oxford Center for Evidence-based Medicine guidelines were used to determine the level of evidence of neurological involvement and treatment based on the reports identified. Results Neurologic manifestations were reported for all NTDs except yaws. Neurologic involvement was described in systematic reviews for four NTDs (Chagas disease, echinococcosis, rabies, cysticercosis) (levels 2a–3a), retrospective cohort studies for six (dengue, human African trypanosomiasis, leishmaniasis, leprosy, onchocerciasis, schistosomiasis) (levels 2b–3b), case series for one (foodborne trematodiasis) (level 4), and case reports for five (Buruli ulcer, dracunculiasis, filariasis, soil-transmitted helminthes, and trachoma). Level 1 evidence for treatment of neurologic manifestations of NTDs was found for human African trypanosomiasis, leprosy, and cysticercosis and level 2 evidence exists for treatment of neurologic involvement in Chagas disease. For the remaining NTDs, treatment of neurologic complications is described in case series and case reports only. Conclusions Neurologic manifestations of NTDs cause significant morbidity and mortality, although limited evidence exists on how best to treat these neurologic complications. Increased awareness of neurologic manifestations of the NTDs can increase their early identification and treatment, contributing to ongoing elimination and eradication campaigns.

The detection of pathogens associated with gastrointestinal disease may be important in certain patient populations, such as immunocompromised hosts, the critically ill, or individuals with prolonged disease that is refractory to treatment. In this study, we evaluated two commercially available multiplex panels (the FilmArray gastrointestinal [GI] panel [BioFire Diagnostics, Salt Lake City, UT] and the Luminex xTag gastrointestinal pathogen panel [GPP] [Luminex Corporation, Toronto, Canada]) using Cary-Blair stool samples ( n = 500) submitted to our laboratory for routine GI testing (e.g., culture, antigen testing, microscopy, and individual real-time PCR). At the time of this study, the prototype (non-FDA-cleared) FilmArray GI panel targeted 23 pathogens (14 bacterial, 5 viral, and 4 parasitic), and testing of 200 μl of Cary-Blair stool was recommended. In contrast, the Luminex GPP assay was FDA cleared for the detection of 11 pathogens (7 bacterial, 2 viral, and 2 parasitic), but had the capacity to identify 4 additional pathogens using a research-use-only protocol. Importantly, the Luminex assay was FDA cleared for 100 μl raw stool; however, 100 μl Cary-Blair stool was tested by the Luminex assay in this study. Among 230 prospectively collected samples, routine testing was positive for one or more GI pathogens in 19 (8.3%) samples, compared to 76 (33.0%) by the FilmArray and 69 (30.3%) by the Luminex assay. Clostridium difficile (12.6 to 13.9% prevalence) and norovirus genogroup I (GI)/GII (5.7 to 13.9% prevalence) were two of the pathogens most commonly detected by both assays among prospective samples. Sapovirus was also commonly detected (5.7% positive rate) by the FilmArray assay. Among 270 additional previously characterized samples, both multiplex panels demonstrated high sensitivity (>90%) for the majority of targets, with the exception of several pathogens, notably Aeromonas sp. (23.8%) by FilmArray and Yersinia enterocolitica (48.1%) by the Luminex assay. Interestingly, the FilmArray and Luminex panels identified mixed infections in 21.1% and 13.0% of positive prospective samples, respectively, compared to only 8.3% by routine methods.

A 65-year-old female with a history of multiple tick bites presented with fever and pancytopenia. Intracytoplasmic rickettsial morulae were detected on peripheral smear and bone marrow biopsy specimens, and PCR amplified Ehrlichia ewingii DNA from both specimens. To our knowledge, this is the first report of E. ewingii infection of human bone marrow.

Objectives:(1) Determine incidence of HPV (human papillomavirus) infection in routine tonsillectomy swabs and specimens. (2) Determine if vaccination impacts tonsillar HPV infection.Methods:This was a prospective study conducted from July 2012 to January 2014. Patients ages 1 to 100 years undergoing tonsillectomy for nonmalignant indications were enrolled. Demographic data and immunization status were collected. Specimens collected during surgery included a flocked swab and half of the left and right tonsil. All specimens were tested for HPV DNA using the polymerase chain reaction–based Roche Cobas Amplicor test and for the presence of E6/E7 mRNA using the Gen-Probe Aptima HPV test.Results:Patients were stratified into 2 groups, pediatric (ages 1-12 years) and adults (age 13 years and up) based on age of potential sexual maturity. In the pediatric group (n = 130) mean age was 5.2 years, 42% were female, 58% were male, and only 3 patients were fully vaccinated against HPV. In the adult group (n = 64) mean ...

An Oregon resident returned from a photography trip to Ethiopia with a male Hyalomma truncatum tick attached to the skin on his lower back. The tick was identified morphologically and deposited in the U.S. National Tick Collection housed at Georgia Southern University, Statesboro, Georgia. The public health importance of Hyalomma species of ticks and diagnostic dilemmas with identifying exotic ticks imported into the U.S. are discussed.

A 65-year-old male with known hypertension and hypercholesterolemia sought medical attention because of a 3-month history of skin swelling on his upper back. Histopathology and molecular techniques were employed and identified an organism in the Onchocerca genus. This represents a very uncommon example of cutaneous infection by a zoonotic Onchocerca species.

SUMMARY The collection, handling, identification, and reporting of ectoparasitic arthropods in clinical and reference diagnostic laboratories are discussed in this review. Included are data on ticks, mites, lice, fleas, myiasis-causing flies, and bed bugs. The public health importance of these organisms is briefly discussed. The focus is on the morphological identification and proper handling and reporting of cases involving arthropod ectoparasites, particularly those encountered in the United States. Other arthropods and other organisms not of public health concern, but routinely submitted to laboratories for identification, are also briefly discussed.

We previously developed a real-time polymerase chain reaction (PCR) assay for detection of the four Plasmodium species that infect humans. Recent studies have shown that natural transmission of the simian parasite Plasmodium knowlesi to humans occurs frequently in Southeast Asia. We have expanded our PCR assay to include detection of P. knowlesi.

Pseudomonas aeruginosa is a frequent and virulent pulmonary pathogen in patients with cystic fibrosis. If colonization is not prevented, P aeruginosa becomes permanently established and nearly always mutates into a mucoid strain. The alginate-containing matrix of the mucoid strain is thought to allow the formation of protected microcolonies and provide increased resistance to opsonization, phagocytosis, and destruction by antibiotics. As a result, conversion to the mucoid phenotype is associated with a significant increase in morbidity and mortality. In the microbiology laboratory, mucoid P aeruginosa has a distinct Gram stain and culture appearance that can expedite its identification and facilitate appropriate patient management. Important aspects of the mucoid phenotype are reviewed.

Diagnosis of yeast infection is typically accomplished by fungal smear and culture, histopathologic examination, and/or serologic studies. Newer assays based on mass spectrometry may be useful for yeast identification when histologic examination is inconclusive, fungal cultures are not ordered, or cultures fail to yield a causative agent. The purpose of this study was to evaluate the ability of the PLEX-ID broad fungal assay to accurately detect and identify yeasts in formalin-fixed paraffin-embedded (FFPE) tissues. Tissue samples from 78 FFPE specimens with both histopathology and corresponding culture results for a variety of yeasts were tested using the PLEX-ID broad fungal assay. A 40-μm FFPE tissue section from each specimen was digested with proteinase K followed by nucleic acid extraction and PCR using broad-range fungal primers. Yeast DNA in amplified products was identified using electrospray ionization mass spectrometry. Discordant results were resolved by D2 rRNA gene sequencing. PLEX-ID analysis detected yeast DNA in 78.2% (61/78) of the cases, of which 91.8% (56/61) were concordant with culture results. Of the 5 discordant positive results, 4 PLEX-ID results were considered to result from environmental contaminants, while 1 clinically important discrepancy was observed ( Blastomyces dermatitidis by culture and Cryptococcus neoformans by PLEX-ID). Sequencing of the discordant sample was unsuccessful. The majority of histopathology results (89.7% [70/78]) correlated with culture results. The PLEX-ID broad fungal assay identifies fungi directly from FFPE tissues and can be a useful adjunct to traditional culture and histopathology tests.

Microsporidia are opportunistic pathogens that usually cause a limited disease in the gastrointestinal tract. Occasionally, they can cause disseminated disease. In solid organ transplant recipients, disseminated disease has been reported only rarely. We describe a 68-year-old woman who presented with fever, cough, and acute kidney injury 6 months after kidney transplantation. Dissemination was confirmed by identification of microsporidial spores in urine and bronchoalveolar lavage fluid. Polymerase chain reaction analysis identified the species as Encephalitozoon cuniculi.

The critical role of the microbiology laboratory in infectious disease diagnosis calls for a close, positive working relationship between the physician and the microbiologists who provide enormous value to the health care team. This document, developed by both laboratory and clinical experts, provides information on which tests are valuable and in which contexts, and on tests that add little or no value for diagnostic decisions. Sections are divided into anatomic systems, including Bloodstream Infections and Infections of the Cardiovascular System, Central Nervous System Infections, Ocular Infections, Soft Tissue Infections of the Head and Neck, Upper Respiratory Infections, Lower Respiratory Tract infections, Infections of the Gastrointestinal Tract, Intraabdominal Infections, Bone and Joint Infections, Urinary Tract Infections, Genital Infections, and Skin and Soft Tissue Infections; or into etiologic agent groups, including Tickborne Infections, Viral Syndromes, and Blood and Tissue Parasite Infections. Each section contains introductory concepts, a summary of key points, and detailed tables that list suspected agents; the most reliable tests to order; the samples (and volumes) to collect in order of preference; specimen transport devices, procedures, times, and temperatures; and detailed notes on specific issues regarding the test methods, such as when tests are likely to require a specialized laboratory or have prolonged turnaround times. There is redundancy among the tables and sections, as many agents and assay choices overlap. The document is intended to serve as a reference to guide physicians in choosing tests that will aid them to diagnose infectious diseases in their patients.

The critical role of the microbiology laboratory in infectious disease diagnosis calls for a close, positive working relationship between the physician and the microbiologists who provide enormous value to the health care team. This document, developed by both laboratory and clinical experts, provides information on which tests are valuable and in which contexts, and on tests that add little or no value for diagnostic decisions. Sections are divided into anatomic systems, including Bloodstream Infections and Infections of the Cardiovascular System, Central Nervous System Infections, Ocular Infections, Soft Tissue Infections of the Head and Neck, Upper Respiratory Infections, Lower Respiratory Tract infections, Infections of the Gastrointestinal Tract, Intraabdominal Infections, Bone and Joint Infections, Urinary Tract Infections, Genital Infections, and Skin and Soft Tissue Infections; or into etiologic agent groups, including Tickborne Infections, Viral Syndromes, and Blood and Tissue Parasite Infections. Each section contains introductory concepts, a summary of key points, and detailed tables that list suspected agents; the most reliable tests to order; the samples (and volumes) to collect in order of preference; specimen transport devices, procedures, times, and temperatures; and detailed notes on specific issues regarding the test methods, such as when tests are likely to require a specialized laboratory or have prolonged turnaround times. There is redundancy among the tables and sections, as many agents and assay choices overlap. The document is intended to serve as a reference to guide physicians in choosing tests that will aid them to diagnose infectious diseases in their patients.

Microbiological diagnosis is pivotal to the appropriate management and treatment of infective endocarditis. We evaluated PCR-electrospray ionization mass spectrometry (PCR/ESI-MS) for bacterial and candidal detection using 83 formalin-fixed paraffin-embedded heart valves from subjects with endocarditis who had positive valve and/or blood cultures, 63 of whom had positive valvular Gram stains. PCR/ESI-MS yielded 55% positivity with concordant microbiology at the genus/species or organism group level (e.g., viridans group streptococci), 11% positivity with discordant microbiology, and 34% with no detection. PCR/ESI-MS detected all antimicrobial resistance encoded by mecA or vanA / B and identified a case of Tropheryma whipplei endocarditis not previously recognized.

Background The gold standard for malaria diagnosis is the examination of thick and thin blood films. Thick films contain 10 to 20 times more blood than thin films, correspondingly providing increased sensitivity for malaria screening. A potential complication of thick film preparations is sloughing of the blood droplet from the slide during staining or rinsing, resulting in the loss of sample. In this work, two methods for improving thick film slide adherence (‘scratch’ (SCM) and ‘acetone dip’ (ADM) methods) were compared to the ‘standard method’ (SM) of thick film preparation. Methods Standardized blood droplets from 26 previously examined EDTA whole blood specimens (22 positive and four negative) were concurrently spread on glass slides using the SM, ADM, and SCM. For the SM and ADM prepared slides, the droplet was gently spread to an approximate 22 millimeters in diameter spot on the slide using the edge of a second glass slide. For the SCM, the droplet was spread by carefully grinding (or scratching) it into the slide with the point of a second glass slide. Slides were dried for one hour in a laminar flow hood. For the ADM, slides were dipped once in an acetone filled Coplin jar and allowed to air dry. All slides were then Giemsa-stained and examined in a blinded manner. Adherence was assessed by blinded reviewers. Results No significant or severe defects were observed for slides prepared with the SCM. In contrast, 8 slides prepared by the ADM and 3 prepared using the SM displayed significant or severe defects. Thick films prepared by the three methods were microscopically indistinguishable and concordant results (positive or negative) were obtained for the three methods. Estimated parasitaemia of the blood samples ranged from 25 to 429,169 parasites/μL of blood. Conclusions The SCM is an inexpensive, rapid, and simple method that improves the adherence of thick blood films to standard glass slides without altering general slide preparation, microscopic appearance or interpretability. Using the SCM, thick films can be reliably examined less than two hours after sample receipt. This represents a significant diagnostic improvement over protocols requiring extended drying periods.

We report the finding of Iodamoeba butschlii amebic cysts on a liquid-based anal Pap smear from an HIV-positive male. Iodine staining of the smear confirmed the diagnosis. It is important to distinguish I. butschlii from pathogenic ameobae and other organisms seen on anal Pap smears.

Testing of clinical plasma specimens by the COBAS AmpliPrep/COBAS TaqMan CMV Test (CAP/CTM CMV), COBAS AMPLICOR CMV MONITOR Test (CAM CMV), and a laboratory-developed assay using analyte-specific reagents (LC CMV) demonstrated substantial result bias for CAP/CTM CMV versus CAM CMV (r = 0.436) and CAP/CTM CMV versus LC CMV (r = 0.773).

We evaluated a commercial multiplex polymerase chain reaction (PCR) assay in a cross-sectional study among 81 adult and pediatric outpatients-40 cases with upper respiratory infection symptoms and 41 asymptomatic controls-from February to April 2008. Two specimens (throat swab and nasal swab) from each participant were tested using the EraGen MultiCode-PLx Respiratory Virus Panel that detects 17 viral targets. Throat swabs were also tested for Group A Streptococcus (GAS) by PCR. Respiratory viruses were detected in 22/40 (55%) cases and in 3/41 (7%) controls (P < 0.001). GAS was detected in 10 (25%) cases; GAS and respiratory virus co-infection was found in 4 (10%). Agreement between nasal and throat swabs for viral detection was 69% in cases and 95% in controls. Of 22 cases with a detectable virus, 12 (54%) were picked up by only 1 (throat or nasal) specimen, and the detection rate was increased by combining results of nasal and throat swab testing.

This article covers the laboratory diagnosis of infections that occur predominantly in the tropics. The discussion includes diagnosis of blood and tissue parasites, intestinal parasites, and tropical infections caused by fungi, bacteria, and mycobacteria. The laboratory performance of techniques for the identification of intestinal parasites and special requirements for the collection of specimens for virology testing are also discussed. Images demonstrating the characteristic features of selected tropical parasites and fungi are included for reference.

This chapter covers protozoa, helminths, and arthropods. 1. Protozoa are single-celled, microscopic eukaryotic organisms like amebae and Giardia. Helminths are parasitic worms including nematodes (roundworms), cestodes (tapeworms), and trematodes (flukes). Arthropods, like ticks and mites, are generally considered parasites. Specific organisms reviewed include Giardia lamblia, Cyclospora cayetanensis, Blastocystis hominis, Entamoeba histolytica, Plasmodium falciparum, Babesia microti, and Toxoplasma gondii. Diagnosis and treatment of different types of infection are also reviewed.

Acute gestational hepatitis from herpes simplex virus (HSV) infection is a rare but potentially life-threatening condition. We present the first reported case of primary HSV type 2 hepatitis in a pregnant woman who was diagnosed by detection of HSV-2 viremia via real-time polymerase chain reaction. The patient was successfully treated with acyclovir and delivered a healthy infant.

Ehrlichiosis is a clinically important, emerging zoonosis. Only Ehrlichia chaffeensis and E. ewingii have been thought to cause ehrlichiosis in humans in the United States. Patients with suspected ehrlichiosis routinely undergo testing to ensure proper diagnosis and to ascertain the cause.We used molecular methods, culturing, and serologic testing to diagnose and ascertain the cause of cases of ehrlichiosis.On testing, four cases of ehrlichiosis in Minnesota or Wisconsin were found not to be from E. chaffeensis or E. ewingii and instead to be caused by a newly discovered ehrlichia species. All patients had fever, malaise, headache, and lymphopenia; three had thrombocytopenia; and two had elevated liver-enzyme levels. All recovered after receiving doxycycline treatment. At least 17 of 697 Ixodes scapularis ticks collected in Minnesota or Wisconsin were positive for the same ehrlichia species on polymerase-chain-reaction testing. Genetic analyses revealed that this new ehrlichia species is closely related to E. muris.We report a new ehrlichia species in Minnesota and Wisconsin and provide supportive clinical, epidemiologic, culture, DNA-sequence, and vector data. Physicians need to be aware of this newly discovered close relative of E. muris to ensure appropriate testing, treatment, and regional surveillance. (Funded by the National Institutes of Health and the Centers for Disease Control and Prevention.).

In June, 2008, a 47-year-old woman presented to us with a 1-day history of crackling sensations in her left ear. On otoscopic examination, a tick was seen on the left tympanic membrane (fi gure). An otorhinolaryngologist removed the tick with endoscopic-guided forceps. There was no bleeding, perforation, hearing loss, or other sequelae. The crackling sensations resolved immediately. The tick was identifi ed as Dermacentor variabilis, commonly known as the American dog tick. Our patient lived in a rural area of Minnesota, USA where the tick-borne diseases lyme and human anaplasmosis are endemic. She denied any known tick exposure, but reported that her dog often slept in her bedroom at night. Fortunately, Dermacentor variabilis is not a primary vector for Borrelia burgdorferi and Anaplasma phagocytophilum, which cause lyme disease and human analasmosis respectively. D variabilis is a known vector of rocky mountain spotted fever and tularaemia, neither of which are common in Minnesota.

We aimed to determine the clinical features, predisposing factors, and outcome of left-sided Pseudomonas aeruginosa endocarditis in persons with no history of injection drug use. We performed a retrospective review of patient medical records from Mayo Clinic (Rochester, MN; Scottsdale, AZ; and Jacksonville, FL) for all cases of left-sided P. aeruginosa endocarditis. We identified 4 cases. We present these cases, as well as a review of the English-language medical literature. Data gathered included the year the case was reported; the valve involved; treatment, including valve replacement surgery; and outcome, if known. Left-sided P. aeruginosa endocarditis in persons without injection drug use is a rare but serious infection, with a history of instrumentation as a common predisposing condition. Valvular surgery is indicated, when possible, for the best chance of survival, along with extended therapy with combination antibiotics for complete recovery.

Pandemic (H1N1) 2009 influenza A virus originated due to triple reassortment of human, avian, and swine viruses (6). The pandemic epicenter was in Mexico, where the first human case was reported on 12 April 2009 (1). Thereafter, 16,813 pandemic H1N1 influenza A-related deaths have been reported to the World Health Organization (WHO) by 213 countries between April 2009 and March 2010 (8). From a laboratory perspective, detection and discrimination of pandemic H1N1 from seasonal and swine H1N1 is crucial for accurate diagnosis and disease surveillance. We previously described a laboratory-developed influenza A virus real-time reverse transcription-PCR (rRT-PCR) assay (Mayo FLU A) for simultaneous identification and subtype discrimination of influenza A virus RNA using melting temperature (Tm) analysis (3, 9). As discussed in the paper by Dhiman et al., loss of subtype discriminatory ability occurred in just 3 months of testing for the pandemic virus, due to multiple point mutations in the targeted 242-base region of the highly conserved matrix (M) gene as determined on a subset of 12 isolates (3). In the current paper, we have extended our analysis to a significantly larger data set to conduct mutational and amino acid analysis and characterize the genetic changes in the M gene of the pandemic H1N1 influenza A virus. Between 1 May 2009 and 31 December 2009, we detected 1,731 influenza A virus clinical isolates out of 9,614 isolates tested (18% positivity rate) using the Mayo FLU A assay; the majority were identified as the pandemic H1N1 subtype based on predefined Tm criteria (50.5°C to 53.2°C). However, between 14 August and 31 December 2009, 48 clinical samples were identified with Tms outside the validated range for pandemic H1N1 (48 of 1,579 isolates [3.0%] during this period), with an overall lower mean Tm of 46.71 ± 1.73°C. Only one specimen, from Ann Arbor, MI, had a higher Tm of 55.5°C, which is unexpected, given that mutations typically cause a decrease rather than an increase in Tm. The significance of this result is unknown, as we did not have additional specimens from that geographical area to confirm if this result was an actual trend or an outlier. All atypical Tm samples were confirmed as pandemic H1N1 using the CDC swine flu assay. We sequenced the 242-base oligonucleotide amplicon in 37 isolates to identify the mutations responsible for the atypical Tms. Mutational analysis revealed multiple point mutations in the 57-bp region detected by the probes compared to the original pandemic H1N1 gene sequence published by the WHO on 28 April 2009 (2) (Table ​(Table1).1). Six of these were silent mutations, resulting in nucleotide changes GAG→GAA, ATC→ATT, GCG→GCA, AGA→AGG, CTG→TTG, and AAC→AAT at amino acid positions 23, 24, 25, 27, 28, and 36, respectively, compared to the original pandemic H1N1 gene sequence. Of interest, two additional nonsynonymous mutations, CTG→ATG and ACA→ATA, resulted in downstream changes in amino acid sequence at positions Leu28Met and Thr37Ile, respectively (Fig. ​(Fig.1).1). We also observed geographic clustering among the mutations in these analyses (Table ​(Table11). FIG. 1. Amino acid sequence of the 57-bp discriminatory region of pandemic (H1N1) 2009 influenza A virus. TABLE 1. Sequence alignment in the discriminatory region of the amplified matrix gene from 37 clinical isolates with atypical Tms using the Mayo FLU A assay Since the emergence of pandemic H1N1, several studies have tried to characterize the mutational trends of this new virus (4, 5, 7). Pan et al. described the emergence of a signature residue at the position of nucleoprotein 100 (NP-100) (valine to isoleucine) that exhibited a dominant change in as many as 93% of isolates by the later phases of the pandemic. In addition, four nonsignature residues, at positions neuraminidase 91 (NA-91), NA-233, hemagglutinin 206 (HA-206), and nonstructural protein 1 (NS1) to NS123, were observed during a short period of time within the epidemic (5). All of these mutant residues were characterized in the viral functional domains, suggesting potent roles in the human adaption and virulence. Mutations were also observed in M1/M2 genes, but they were of low frequency with an unknown role. Potdar et al. reported several mutations throughout the viral genome, including a predominant D225G mutation in the H gene present in the receptor binding domain of Indian novel H1N1 isolates (7). Nelson et al. conducted a whole-genome phylogenetic analysis on 290 isolates of pandemic H1N1 influenza virus collected globally and identified seven major clades that have cocirculated worldwide since April 2009 (4). Several amino acid changes were also identified, predominantly in the HA, NA, and NP genes. However, no mutations were observed in the matrix gene in the latter two studies. In our study, we found eight mutations in a very short sequence span of only 57 bp of the influenza A virus matrix gene, of which two were nonsynonymous (Leu28Met and Thr37Ile). This is an unexpected finding, as the influenza A virus matrix gene is relatively conserved and is not thought to be under the same genetic selective pressures as other genes, such as HA and NA, that have very high mutability rates. The implications of these findings for viral adaptation and virulence have yet to be determined. However, from a diagnostic perspective, these mutations resulted in the loss of viral subtype discriminatory ability using Tm analysis within just 3 months of the pandemic and hindered laboratory diagnosis. Evidence for mutability in the highly conserved M gene of influenza virus calls for pandemic as well as routine in silico monitoring of primers and probes for optimal target coverage.