Pathology and Monkeypox virus Localization in Tissues From Immunocompromised Patients With Severe or Fatal Mpox.

BACKGROUND: Pathology and Monkeypox virus (MPXV) tissue tropism in severe and fatal human mpox is not thoroughly described but can help elucidate the disease pathogenesis and the role of coinfections in immunocompromised patients. METHODS: We analyzed biopsy and autopsy tissues from 22 patients with severe or fatal outcomes to characterize pathology and viral antigen and DNA distribution in tissues by immunohistochemistry and in situ hybridization. Tissue-based testing for coinfections was also performed. RESULTS: Mucocutaneous lesions showed necrotizing and proliferative epithelial changes. Deceased patients with autopsy tissues evaluated had digestive tract lesions, and half had systemic tissue necrosis with thrombotic vasculopathy in lymphoid tissues, lung, or other solid organs. Half also had bronchopneumonia, and one-third had acute lung injury. All cases had MPXV antigen and DNA detected in tissues. Coinfections were identified in 5 of 16 (31%) biopsy and 4 of 6 (67%) autopsy cases. CONCLUSIONS: Severe mpox in immunocompromised patients is characterized by extensive viral infection of tissues and viremic dissemination that can progress despite available therapeutics. Digestive tract and lung involvement are common and associated with prominent histopathological and clinical manifestations. Coinfections may complicate mpox diagnosis and treatment. Significant viral DNA (likely correlating to infectious virus) in tissues necessitates enhanced biosafety measures in healthcare and autopsy settings.

Rickettsia honei Infection in a Traveler Returning From India.

We report a case of Rickettsia honei infection in a US tourist returning from India and the Himalayas. This case highlights a need for awareness of various Rickettsia species endemic to India and the importance for physicians to consider rickettsial diseases in returning travelers with eschar or rash-associated febrile illnesses.

Diagnosis of Spotted Fever Group Rickettsioses in U.S. Travelers Returning from Africa, 2007-2016.

Spotted fever group rickettsioses (SFGRs), such as African tick bite fever (ATBF), are among the most commonly diagnosed diseases for ill travelers returning from southern Africa. We summarized demographic, clinical, and diagnostic features of imported SFGR cases in U.S. travelers returning from Africa who had laboratory specimens submitted to the Centers for Disease Control and Prevention. Diagnosis of SFGR was performed by indirect immunofluorescence antibody assay, immunohistochemical staining, polymerase chain reaction (PCR), or culture. Cases were defined as probable SFGR, confirmed SFGR, or confirmed ATBF. Clinical and epidemiological categorical variables were described as counts and proportions; continuous variables were described using geometric mean titers, median, and range. One hundred and twenty-seven patients satisfied laboratory criteria for confirmed or probable SFGR. Fever was the most common symptom (N = 88; 69%), followed by ≥ 1 eschars (N = 70; 55%). Paired serums were submitted for 36 patients (28%); 12 patients (33%) had nonreactive initial serum sample but converted to a titer ≥ 64 with the convalescent sample. Twenty-seven patients (21%) had infection with Rickettsia africae based on PCR analysis of eschar swab (N = 8) or biopsy (N = 23). Fifteen patients had eschar biopsy or swab samples and serum sample(s) submitted together; 9 (60%) had PCR-positive eschar results and nonreactive acute serology. Health-care providers should consider SFGR when evaluating patients for a febrile illness with eschar and compatible foreign travel history. Polymerase chain reaction testing of eschar biopsies or swabs provides a confirmed diagnosis in early stages of disease; eschar swabs or biopsies are an underutilized diagnostic technique.

Ebola Virus Disease in Pregnancy: Clinical, Histopathologic, and Immunohistochemical Findings.

Here we describe clinicopathologic features of Ebola virus disease in pregnancy. One woman infected with Sudan virus in Gulu, Uganda, in 2000 had a stillbirth and survived, and another woman infected with Bundibugyo virus had a live birth with maternal and infant death in Isiro, the Democratic Republic of the Congo in 2012. Ebolavirus antigen was seen in the syncytiotrophoblast and placental maternal mononuclear cells by immunohistochemical analysis, and no antigen was seen in fetal placental stromal cells or fetal organs. In the Gulu case, ebolavirus antigen localized to malarial parasite pigment-laden macrophages. These data suggest that trophoblast infection may be a mechanism of transplacental ebolavirus transmission.

Notes from the Field: Rickettsia parkeri Rickettsiosis - Georgia, 2012-2014.

During 2012-2014, five cases of Rickettsia parkeri rickettsiosis were identified by a single urgent care practice in Georgia, located approximately 40 miles southwest of Atlanta. Symptom onset occurred during June-October, and all patients had a known tick bite. Patients ranged in age from 27 to 72 years (median = 53 years), and all were male. The most commonly reported initial signs were erythema (n = 3) and swelling (n = 2) at the site of the bite. Two patients reported fever and a third patient reported a rash and lymphadenopathy without fever. Other symptoms included myalgia (n = 3), chills (n = 3), fatigue (n = 2), arthralgia (n = 2), and headache (n = 2). Eschar biopsy specimens were collected from each patient using a 4-mm or 5-mm punch and placed in 10% neutral buffered formalin or sterile saline. These specimens were tested by immunohistochemical (IHC) stains, quantitative polymerase chain reaction (qPCR) assays, or cell culture isolation to determine if there was evidence of infection with a Rickettsia species (1). IHC evidence of spotted fever group rickettsiae was found in the eschar biopsy specimens in all five cases. In four cases, the biopsy specimens were also positive for R. parkeri by qPCR. The fifth case (specimen positive only by IHC testing) was considered a probable R. parkeri case based on clinical signs and symptoms. R. parkeri was grown in cell culture from one specimen from which isolation was attempted. All patients were treated with oral doxycycline (100 mg twice daily) for a minimum of 10 days, and all recovered.

Immunolocalization and Distribution of Rubella Antigen in Fatal Congenital Rubella Syndrome.

BACKGROUND: An estimated 100,000 cases of congenital rubella syndrome (CRS) occur worldwide each year. The reported mortality rate for infants with CRS is up to 33%. The cellular mechanisms responsible for the multiple congenital defects in CRS are presently unknown. Here we identify cell types positive for rubella virus (RV) in CRS infants. METHODS: Cells and organs involved in RV replication were identified in paraffin-embedded autopsy tissues from three fatal case-patients by histopathologic examination and immunohistochemical (IHC) staining using a rabbit polyclonal RV antibody. Normal rabbit antisera and RV antisera preabsorbed with highly purified RV served as negative controls. RESULTS: RV antigen was found in interstitial fibroblasts in the heart, adventitial fibroblasts of large blood vessels, alveolar macrophages, progenitor cells of the outer granular layer of the brain, and in capillary endothelium and basal plate in the placenta. The antibody specificity was verified by IHC staining of multiple tissue sections from other infectious disease cases. RV infection of each cell type is consistent with abnormalities which have been identified in patients with CRS, in the heart, large blood vessels, and brain. Antigen distribution was consistent with inflammatory response to vascular injury and systemic spread of RV. CONCLUSIONS: The identification of RV positive cell types in CRS is important to better understand the pathology and pathogenesis of CRS.

Immunoproteomic profiling of Rickettsia parkeri and Rickettsia amblyommii.

Rickettsia parkeri is an Amblyomma-associated, spotted fever group Rickettsia species that causes an eschar-associated, febrile illness in multiple countries throughout the Western Hemisphere. Many other rickettsial species of known or uncertain pathogenicity have been detected in Amblyomma spp. ticks in the Americas, including Rickettsia amblyommii, "Candidatus Rickettsia andeanae" and Rickettsia rickettsii. In this study, we utilized an immunoproteomic approach to compare antigenic profiles of low-passage isolates of R. parkeri and R. amblyommii with serum specimens from patients with PCR- and culture-confirmed infections with R. parkeri. Five immunoreactive proteins of R. amblyommii and nine immunoreactive proteins of R. parkeri were identified by matrix-assisted laser desorption ionization tandem time-of-flight mass spectrometry. Four of these, including the outer membrane protein (Omp) A, OmpB, translation initiation factor IF-2, and cell division protein FtsZ, were antigens common to both rickettsiae. Serum specimens from patients with R. parkeri rickettsiosis reacted specifically with cysteinyl-tRNA synthetase, DNA-directed RNA polymerase subunit alpha, putative sigma (54) modulation protein, chaperonin GroEL, and elongation factor Tu of R. parkeri which have been reported as virulence factors in other bacterial species. Unique antigens identified in this study may be useful for further development of the better serological assays for diagnosing infection caused by R. parkeri.

First reported case of Ehrlichia ewingii involving human bone marrow.

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.

Detection of Rickettsia rickettsii, Rickettsia parkeri, and Rickettsia akari in skin biopsy specimens using a multiplex real-time polymerase chain reaction assay.

BACKGROUND: Rickettsia rickettsii, Rickettsia parkeri, and Rickettsia akari are the most common causes of spotted fever group rickettsioses indigenous to the United States. Infected patients characteristically present with a maculopapular rash, often accompanied by an inoculation eschar. Skin biopsy specimens are often obtained from these lesions for diagnostic evaluation. However, a species-specific diagnosis is achieved infrequently from pathologic specimens because immunohistochemical stains do not differentiate among the causative agents of spotted fever group rickettsiae, and existing polymerase chain reaction (PCR) assays generally target large gene segments that may be difficult or impossible to obtain from formalin-fixed tissues. METHODS: This work describes the development and evaluation of a multiplex real-time PCR assay for the detection of these 3 Rickettsia species from formalin-fixed, paraffin-embedded (FFPE) skin biopsy specimens. RESULTS: The multiplex PCR assay was specific at discriminating each species from FFPE controls of unrelated bacterial, viral, protozoan, and fungal pathogens that cause skin lesions, as well as other closely related spotted fever group Rickettsia species. CONCLUSIONS: This multiplex real-time PCR demonstrates greater sensitivity than nested PCR assays in FFPE tissues and provides an effective method to specifically identify cases of Rocky Mountain spotted fever, rickettsialpox, and R. parkeri rickettsiosis by using skin biopsy specimens.

Microsporidiosis acquired through solid organ transplantation: a public health investigation.

BACKGROUND: Encephalitozoon cuniculi, a microsporidial species most commonly recognized as a cause of renal, respiratory, and central nervous system infections in immunosuppressed patients, was identified as the cause of a temporally associated cluster of febrile illness among 3 solid organ transplant recipients from a common donor. OBJECTIVE: To confirm the source of the illness, assess donor and recipient risk factors, and provide therapy recommendations for ill recipients. DESIGN: Public health investigation. SETTING: Two transplant hospitals and community interview with the deceased donor's family. PATIENTS: Three transplant recipients and the organ donor. MEASUREMENTS: Specimens were tested for microsporidia by using culture, immunofluorescent antibody, polymerase chain reaction,immunohistochemistry, and electron microscopy. Donor medical records were reviewed and a questionnaire was developed to assess for microsporidial infection. RESULTS: Kidneys and lungs were procured from the deceased donor and transplanted to 3 recipients who became ill with fever 7 to 10 weeks after the transplant. Results of urine culture, serologic,and polymerase chain reaction testing were positive for E. cuniculi of genotype III in each recipient; the organism was also identified in biopsy or autopsy specimens in all recipients. The donor had positive serologic test results for E. cuniculi. Surviving recipients received albendazole. Donor assessment did not identify factors for suspected E. cuniculi infection. LIMITATION: Inability to detect organism by culture or polymerase chain reaction in donor due to lack of autopsy specimens. CONCLUSION: Microsporidiosis is now recognized as an emerging transplant-associated disease and should be considered in febrile transplant recipients when tests for routinely encountered agents are unrevealing. Donor-derived disease is critical to assess when multiple recipients from a common donor are ill.

Exserohilum infections associated with contaminated steroid injections: a clinicopathologic review of 40 cases.

September 2012 marked the beginning of the largest reported outbreak of infections associated with epidural and intra-articular injections. Contamination of methylprednisolone acetate with the black mold, Exserohilum rostratum, was the primary cause of the outbreak, with >13,000 persons exposed to the potentially contaminated drug, 741 confirmed drug-related infections, and 55 deaths. Fatal meningitis and localized epidural, paraspinal, and peripheral joint infections occurred. Tissues from 40 laboratory-confirmed cases representing these various clinical entities were evaluated by histopathological analysis, special stains, and IHC to characterize the pathological features and investigate the pathogenesis of infection, and to evaluate methods for detection of Exserohilum in formalin-fixed, paraffin-embedded (FFPE) tissues. Fatal cases had necrosuppurative to granulomatous meningitis and vasculitis, with thrombi and abundant angioinvasive fungi, with extensive involvement of the basilar arterial circulation of the brain. IHC was a highly sensitive method for detection of fungus in FFPE tissues, demonstrating both hyphal forms and granular fungal antigens, and PCR identified Exserohilum in FFPE and fresh tissues. Our findings suggest a pathogenesis for meningitis involving fungal penetration into the cerebrospinal fluid at the injection site, with transport through cerebrospinal fluid to the basal cisterns and subsequent invasion of the basilar arteries. Further studies are needed to characterize Exserohilum and investigate the potential effects of underlying host factors and steroid administration on the pathogenesis of infection.

Gastrointestinal basidiobolomycosis treated with posaconazole.

A 67 year-old Caucasian male from Arizona presented with indolent symptoms of intestinal obstruction and hydronephrosis, found at surgery to be caused by a mass involving the terminal ileum and cecum, extending into the posterior abdominal wall and obstructing the right ureter. Histopathology was diagnostic of basidiobolomycosis. PCR of tissue and sequencing identified the fungus as, Basidiobolus ranarum. During one year of posaconazole treatment, the residual mass shrank, hydronephrosis was relieved and peripheral eosinophilia resolved.

Ultrastructural characterization of pandemic (H1N1) 2009 virus.

We evaluated pandemic influenza A (H1N1) 2009 virus isolates and respiratory tissues collected at autopsy by electron microscopy. Many morphologic characteristics were similar to those previously described for influenza virus. One of the distinctive features was dense tubular structures in the nuclei of infected cells.

Fatal Cokeromyces recurvatus pneumonia: report of a case highlighting the potential for histopathologic misdiagnosis as coccidioides.

Cokeromyces recurvatus is a dimorphic zygomycete with histologic morphology similar to Coccidioides immitis. A 66-year-old man who was status-post bone marrow transplantation for chronic myelogenous leukemia was hospitalized with new onset rash, nausea, and vomiting and subsequently expired. A sputum culture collected on the day of death revealed heavy growth of C. recurvatus 6 days after collection. At autopsy, microscopic examination of the lungs revealed numerous thick-walled, nonbudding spherules ranging in size from 40 to 80 µm. Initial immunohistochemical staining of the formalin-fixed lung tissue was positive for Coccidioides. Additional immunoperoxidase staining revealed the organisms were consistent with a zygomycete fungus, compatible with C. recurvatus infection. Polymerase chain reaction using panfungal primers was attempted on the formalin-fixed tissue but was inconclusive. This case highlights the potential for misdiagnosing Cokeromyces as Coccidioides when the diagnosis is based on histology and immunohistochemical staining.

The expanding spectrum of eschar-associated rickettsioses in the United States.

BACKGROUND: Until recently, Rickettsia rickettsii was the only substantiated cause of tick-borne spotted fever group (SFG) rickettsiosis in humans in the United States. Rickettsia parkeri, originally thought to be nonpathogenic in humans, was recently proved to be another cause of tick-borne SFG rickettsiosis. OBSERVATIONS: We report 3 cases of SFG rickettsiosis and discuss the epidemiology, clinical presentation, histopathologic features, and laboratory findings that support confirmed or probable diagnoses of R parkeri infection and describe the expanding list of eschar-associated SFG rickettsioses recognized in US patients. CONCLUSIONS: The SFG rickettsioses share many clinical manifestations and extensive antigenic cross-reactivity that may hamper specific confirmation of the causative agent.

Rickettsia 364D: a newly recognized cause of eschar-associated illness in California.

BACKGROUND: Four spotted fever group rickettsiae (SFGR) are known to infect humans in the United States. A member of the SFGR designated 364D and detected in Dermacentor occidentalis ticks has not previously been identified as a human pathogen. METHODS: An 80-year-old man from a rural northern California community presented with an eschar on his forearm. A skin punch biopsy of the lesion was evaluated by immunohistochemistry and molecular analysis. Serum specimens obtained from the patient and 3 other area residents with similar illnesses were tested by immunofluorescence and Western immunoblot for antibodies to SFGR. Ticks were collected near the patient's residence and tested for SFGR. RESULTS: Abundant intracellular rickettsiae and fragmented rickettsial antigens were observed in the mononuclear inflammatory infiltrates of the biopsy. Nucleotide sequences of DNA fragments amplified from the biopsy were identical to those of 364D. Convalescent sera from all four patients exhibited high immunoglobulin G titers to Rickettsia rickettsii, Rickettsia rhipicephali, and 364D antigens. Three adult D. occidentalis were positive for 364D, R. rhipicephali, and an unidentified Rickettsia species. CONCLUSIONS: This is the first confirmation of human disease associated with the SFGR 364D, which was likely transmitted by D. occidentalis. Although the patients described here presented with a single cutaneous eschar as the principal manifestation, the full spectrum of illness associated with 364D has yet to be determined. Possible infection with 364D or other SFGR should be confirmed through molecular techniques in patients who present with "spotless" Rocky Mountain spotted fever or have serum antibodies to R. rickettsii with group-specific assays.

Cellular immune responses to severe acute respiratory syndrome coronavirus (SARS-CoV) infection in senescent BALB/c mice: CD4+ T cells are important in control of SARS-CoV infection.

We characterized the cellular immune response to severe acute respiratory syndrome coronavirus (SARS-CoV) infection in 12- to 14-month-old BALB/c mice, a model that mimics features of the human disease. Following intranasal administration, the virus replicated in the lungs, with peak titers on day 2 postinfection. Enhanced production of cytokines (tumor necrosis factor alpha [TNF-alpha] and interleukin-6 [IL-6]) and chemokines (CXCL10, CCL2, CCL3, and CCL5) correlated with migration of NK cells, macrophages, and plasmacytoid dendritic cells (pDC) into the lungs. By day 7, histopathologic evidence of pneumonitis was seen in the lungs when viral clearance occurred. At this time, a second wave of enhanced production of cytokines (TNF-alpha, IL-6, gamma interferon [IFN-gamma], IL-2, and IL-5), chemokines (CXCL9, CXCL10, CCL2, CCL3, and CCL5), and receptors (CXCR3, CCR2, and CCR5), was detected in the lungs, associated with an influx of T lymphocytes. Depletion of CD8(+) T cells at the time of infection did not affect viral replication or clearance. However, depletion of CD4(+) T cells resulted in an enhanced immune-mediated interstitial pneumonitis and delayed clearance of SARS-CoV from the lungs, which was associated with reduced neutralizing antibody and cytokine production and reduced pulmonary recruitment of lymphocytes. Innate defense mechanisms are able to control SARS-CoV infection in the absence of CD4(+) and CD8(+) T cells and antibodies. Our findings provide new insights into the pathogenesis of SARS, demonstrating the important role of CD4(+) but not CD8(+) T cells in primary SARS-CoV infection in this model.

Proteomic analysis of Rickettsia parkeri strain portsmouth.

Rickettsia parkeri, a recently recognized pathogen of human, is one of several Rickettsia spp. in the United States that causes a spotted fever rickettsiosis. To gain insights into its biology and pathogenesis, we applied the proteomics approach to establish a two-dimensional gel proteome reference map and combined this technique with cell surface biotinylation to identify surface-exposed proteins of a low-passage isolate of R. parkeri obtained from a patient. We identified 91 proteins by matrix-assisted laser desorption ionization-tandem time of flight mass spectrometry. Of these, 28 were characterized as surface proteins, including virulence-related proteins (e.g., outer membrane protein A [OmpA], OmpB, beta-peptide, and RickA). Two-dimensional immunoblotting with serum from the R. parkeri-infected index patient was utilized to identify the immunoreactive proteins as potential targets for diagnosis and vaccine development. In addition to the known rickettsial antigens, OmpA and OmpB, we identified translation initiation factor 2, cell division protein FtsZ, and cysteinyl-tRNA synthetase as immunoreactive proteins. The proteome map with corresponding cell surface protein analysis and antigen detection will facilitate a better understanding of the mechanisms of rickettsial pathogenesis.

Risk factors for lower respiratory tract infection death among infants in the United States, 1999-2004.

OBJECTIVE: To describe maternal and birth-related risk factors associated with lower respiratory tract infection (LRTI) deaths among infants. METHODS: Records for infants with LRTI as a cause of death were examined by using the linked birth/infant death database for 1999-2004. Singleton infants dying with LRTI and a random sample of surviving singleton infants were compared for selected characteristics. RESULTS: A total of 5420 LRTI-associated infant deaths were documented in the United States during 1999-2004, for an LRTI-associated infant mortality rate of 22.3 per 100,000 live births. Rates varied according to race; the rate for American Indian/Alaska Native (AI/AN) infants was highest (53.2), followed by black (44.1), white (18.7), and Asian/Pacific Islander infants (12.3). Singleton infants with low birth weight (<2500 g) were at increased risk of dying with LRTI after controlling for other characteristics, especially black infants. Both AI/AN and black infants born with a birth weight of > or =2500 g were more likely to have died with LRTI than other infants of the same birth weight. Other risk factors associated with LRTI infant death included male gender, the third or more live birth, an Apgar score of <8, unmarried mother, mother with <12 years of education, mother <25 years of age, and mother using tobacco during pregnancy. CONCLUSIONS: Low birth weight was associated with markedly increased risk for LRTI-associated death among all of the racial groups. Among infants with a birth weight of > or =2500 g, AI/AN and black infants were at higher risk of LRTI-associated death, even after controlling for maternal and birth-related factors. Additional studies and strategies should focus on the prevention of maternal and birth-related risk factors for postneonatal LRTI and on identifying additional risk factors that contribute to elevated mortality among AI/AN and black infants.