Prevalence of genotoxic bacteria in men undergoing biopsy for prostate cancer.

BACKGROUND: New evidence suggests that bacteria-produced DNA toxins may have a role in the development or progression of prostate cancer. To determine the prevalence of these genes in a noninfection (i.e., colonized) state, we screened urine specimens in men before undergoing a biopsy for prostate cancer detection. METHODS: We developed a multiplex polymerase chain reaction using three of the most described bacterial genotoxin gene primers: Colibactin (polyketone synthase [pks] gene island: clbN and clbB), cytotoxic necrotizing factor (cnf1) toxin, and cytolethal distending toxin B (cdtB) represented gene islands. After calibration on Escherichia coli samples of known genotypes, we used a training and validation cohort. We performed multiplex testing on a training cohort of previously collected urine from 45 men undergoing prostate biopsy. For the validation cohort, we utilized baseline urine samples from a previous randomized clinical trial (n = 263) with known prostate cancer outcomes. RESULTS: The prevalence of four common bacterial genotoxin genes detected in the urine before prostate biopsy for prostate cancer is 8% (25/311). The prevalence of pks island (clbN and clbB), cnf1, and cdt toxin genes are 6.1%, 2.4%, and 1.7%, respectively. We found no association between urinary genotoxins and prostate cancer (p = 0.83). We did identify a higher proportion of low-grade cancer (92% vs. 44%) in those men positive for urinary genotoxin and higher-grade cancer in those genotoxin negative (8% vs. 56%, p = 0.001). CONCLUSIONS: The prevalence of urinary genotoxins is low and does not correspond to a prostate cancer diagnosis. The urine was taken at one point in time and does not rule out the possibility of previous exposure.

Molecular genetic testing does not improve the detection of fluoroquinolone resistance before transrectal prostate biopsy.

Background: Fluoroquinolone-resistant (FQR) Escherichia coli (E. coli) causes transrectal prostate biopsy infections. We seek to further identify fluoroquinolones resistance by the incorporation of genetic profiling to influence antibiotic selection for transrectal prostate biopsy and whether the addition of this genetic testing could improve the prediction of FQR detection at the time of biopsy. Materials and methods: In this prospective observational cohort study, rectal swabs were collected within 30 days of an upcoming prostate biopsy. These swabs were sent for phenotypic and genotypic assessment to predict FQR on the day of the biopsy. Phenotype: Specimens were inoculated onto MacConkey agar containing ciprofloxacin using standard culture techniques to determine FQR status. Genotype: We compared cultures to polymerase chain reaction (PCR) sequence typing (E.coli- ST131/H30/ST69) and bacterial plasmids (gyrA, qnrQ, and qnrS). The presence of FQR on this testing was compared to the second rectal swab collected just before biopsy (2 hours after ciprofloxacin prophylaxis), which served as the gold standard for FQR. Results: Overall, the FQR rate was 23.6%. The bacterial plasmids (qnr) were present in 54.1% of samples, and multidrug-resistant E. coli ST131 was present in 12.5% of samples. In comparison, phenotypic assessment using rectal culture had a better prediction for the presence of FQR as compared to genotypic testing [area under the curve (AUC) = 0.85 in phenotype arm vs. AUC = 0.45 in genotype arm]. Conclusion: We detected a high prevalence of FQR genes in the rectum, but the addition of PCR-based genotyping did not improve the prediction of culture-based FQR at the time of biopsy.

Phaeohyphomycosis due to Veronaea botryosa in cultured white sturgeon (Acipenser transmontanus Richardson) from California USA during 2006 to 2015.

Infection with Veronaea botryosa can result in rare cutaneous or disseminated, granulomatous to pyogranulomatous phaeohyphomycosis in humans, although disease due to the fungus has also been reported in non-mammalian vertebrates. This report documents disease due to V. botryosa in captive, juvenile to subadult or young adult white sturgeon (Acipenser transmontanus Richardson) from California USA and complements a previous report of the disease in captive Siberian sturgeon (Acipenser baerii) from Florida USA. Pathological examinations revealed granulomatous to pyogranulomatous inflammation of multiple organs. Isolates of the fungal agent were phenotypically consistent with V. botryosa, and molecular analyses of the D1/D2 region of the fungal 28S rRNA gene and the internal transcribed spacer (ITS) region located between the fungal 18S and 28S rRNA genes confirmed the aetiologic agent as V. botryosa. The disease in captive sturgeon results in a considerable economic encumbrance to the producer due to the loss of the cumulative financial resources invested in the production of older subadult to young adult sturgeon.

Genotyping of Cytomegalovirus from Symptomatic Infected Neonates in Iraq.

Among all other viruses, human cytomegalovirus (HCMV) is the most frequent cause of congenital infection worldwide. Strain variation in HCMV may predict severity or outcome of congenital HCMV disease. Previous studies have associated a particular genotype with specific sequelae or more severe illness, but the results were contradictory. There are no previous studies addressing the genotype of HCMV in Iraq. Therefore, the present study is aimed at molecular detection and genotyping of HCMV isolated from symptomatic congenitally/perinatally infected neonates. This prospective study comprised 24 serum samples from symptomatic neonates with congenital/perinatal infection. Viral DNA was extracted from these serum samples; nested polymerase chain reaction was used to amplify the HCMV gB (UL55) gene. Polymerase chain reaction products of the second round of amplification were subjected to direct Sanger sequencing. Bioedit and MEGA5 software (EMBL-EBI, Hinxton, Cambridgeshire, UK) were used for alignment and construction of a phylogenetic tree. Human cytomegalovirus DNA was detected in 23 of 24 samples (95.8%). According to the phylogenetic analysis, three genotypes of the virus were identified; gB1, gB2, and gB3 genotypes. However, the gB4 genotype was not detected. Human cytomegalovirus gB3 was the most frequent genotype: 14 of 24 (58.33%) among symptomatic infected infants, followed by gB1 (6/24; 25%) and gB2 (4/24; 16.67%). A mixed HCMV infection with gB3/gB1 was detected in only one case. Human cytomegalovirus gB3 was the most predominant genotype among symptomatic congenitally/perinatally HCMV-infected neonates. No association was found between B3 genotype and specific clinical presentation. Jaundice was the most common clinical feature among symptomatically infected neonates, followed by hepatosplenomegaly.

Fatal disseminated Rasamsonia infection in cystic fibrosis post-lung transplantation.

BACKGROUND: Disseminated fungal infections are a known serious complication in individuals with cystic fibrosis (CF) following orthotopic lung transplantation. Aspergillus fumigatus and Scedosporium species are among the more common causes of invasive fungal infection in this population. However, it is also important for clinicians to be aware of other emerging fungal species which may require markedly different antifungal therapies. CASE SUMMARY: We describe the first laboratory-documented case of a fatal disseminated fungal infection caused by Rasamsonia aegroticola in a 21-year-old female CF patient status post-bilateral lung transplantation, which was only identified post-mortem. Molecular analysis revealed the presence of the identical Rasamsonia strains in the patient's respiratory cultures preceding transplantation. DISCUSSION: We propose that the patient's disseminated fungal disease and death occurred as a result of recrudescence of Rasamsonia infection from her native respiratory system in the setting of profound immunosuppression post-operatively. Since Rasamsonia species have been increasingly recovered from the respiratory tract of CF patients, we further review the literature on these fungi and discuss their association with invasive fungal infections in the CF lung transplant host. CONCLUSION: Our report suggests Rasamsonia species may be important fungal pathogens that may have fatal consequences in immunosuppressed CF patients after solid organ transplantation.

New disease records for hatchery-reared sturgeon. II. Phaeohyphomycosis due to Veronaea botryosa.

A series of fungal cases in hatchery-reared juvenile and young adult Siberian sturgeon Acipenser baerii and white sturgeon A. transmontanus occurred at production facilities in Florida and California, USA, respectively. Affected fish exhibited abnormal orientation and/or buoyancy, emaciation, coelomic distension, exophthalmos, cutaneous erythema, and ulcerative skin and eye lesions. Necropsies revealed haemorrhage throughout the coelom, serosanguinous coelomic effusion and organomegaly with nodular or cystic lesions in multiple organs. Fungal hyphae were observed in 27 fish (24 A. baerii and 3 A. transmontanus) via microscopic examination of tissue wet mounts and on slides prepared from colonies grown on culture media. Histopathological examination of these infected tissues revealed extensive infiltration by melanised fungal hyphae that were recovered in culture. Phenotypic characteristics and sequencing of the fungal isolates with the use of the internal transcribed spacer region and 28S rRNA gene confirmed the aetiological agent as Veronaea botryosa. To our knowledge, this is the first documentation of V. botryosa infection in fish, although melanised fungi of the closely related genus Exophiala are well-known pathogens of freshwater and marine fishes.

Paravertebral mushroom: identification of a novel species of Phellinus as a human pathogen in chronic granulomatous disease.

We describe a case of paravertebral abscess caused by a Phellinus sp. in a boy with chronic granulomatous disease. Sequence-based identification of this mold, a new agent of disease, suggests a close relation to Phellinus umbrinellus.

Identifying DNA mutations in purified hematopoietic stem/progenitor cells.

In recent years, it has become apparent that genomic instability is tightly related to many developmental disorders, cancers, and aging. Given that stem cells are responsible for ensuring tissue homeostasis and repair throughout life, it is reasonable to hypothesize that the stem cell population is critical for preserving genomic integrity of tissues. Therefore, significant interest has arisen in assessing the impact of endogenous and environmental factors on genomic integrity in stem cells and their progeny, aiming to understand the etiology of stem-cell based diseases. LacI transgenic mice carry a recoverable λ phage vector encoding the LacI reporter system, in which the LacI gene serves as the mutation reporter. The result of a mutated LacI gene is the production of ß-galactosidase that cleaves a chromogenic substrate, turning it blue. The LacI reporter system is carried in all cells, including stem/progenitor cells and can easily be recovered and used to subsequently infect E. coli. After incubating infected E. coli on agarose that contains the correct substrate, plaques can be scored; blue plaques indicate a mutant LacI gene, while clear plaques harbor wild-type. The frequency of blue (among clear) plaques indicates the mutant frequency in the original cell population the DNA was extracted from. Sequencing the mutant LacI gene will show the location of the mutations in the gene and the type of mutation. The LacI transgenic mouse model is well-established as an in vivo mutagenesis assay. Moreover, the mice and the reagents for the assay are commercially available. Here we describe in detail how this model can be adapted to measure the frequency of spontaneously occurring DNA mutants in stem cell-enriched Lin(-)IL7R(-)Sca-1(+)cKit(++)(LSK) cells and other subpopulations of the hematopoietic system.

Invasive fungal infection due to Triadelphia pulvinata in a patient with acute myeloid leukemia.

Triadelphia pulvinata is a rare dematiaceous fungus found in soil. We report the first case of invasive disease in a patient with acute myeloid leukemia who had a bloodstream infection with possibly both lung and brain involvement. Identification was by combined phenotypic features and fungal ribosomal DNA sequence analysis.

The production of monokaryotic hyphae by Cryptococcus neoformans can be induced by high temperature arrest of the cell cycle and is independent of same-sex mating.

Cryptococcus neoformans is a heterothallic fungal pathogen of humans and animals. Although the fungus grows primarily as a yeast, hyphae are produced during the sexual phase and during a process called monokaryotic fruiting, which is also believed to involve sexual reproduction, but between cells of the same mating type. Here we report a novel monokaryotic fruiting mechanism that is dependent on the cell cycle and occurs in haploid cells in the absence of sexual reproduction. Cells grown at 37°C were found to rapidly produce hyphae (∼4 hrs) and at high frequency (∼40% of the population) after inoculation onto hyphae-inducing agar. Microscopic examination of the 37°C seed culture revealed a mixture of normal-sized and enlarged cells. Micromanipulation of single cells demonstrated that only enlarged cells were able to produce hyphae and genetic analysis confirmed that hyphae did not arise from α-α mating or endoduplication. Cell cycle analysis revealed that cells grown at 37°C had an increased population of cells in G2 arrest, with the proportion correlated with the frequency of monokaryotic fruiting. Cell sorting experiments demonstrated that enlarged cells were only found in the G2-arrested population and only this population contained cells able to produce hyphae. Treatment of cells at low temperature with the G2 cell cycle arrest agent, nocodazole, induced hyphal growth, confirming the role of the cell cycle in this process. Taken together, these results reveal a mating-independent mechanism for monokaryotic fruiting, which is dependent on the cell cycle for induction of hyphal competency.

Mixed infection caused by Lecythophora canina sp. nov. and Plectosphaerella cucumerina in a German shepherd dog.

We describe an opportunistic, disseminated infection in a German shepherd dog associated with two fungal organisms not previously reported to cause disease. Lecythophora canina, a new species here described, was isolated from an osteolytic bone lesion. A fine needle aspirate of the lesion demonstrated septate hyphae. Plectospharella cucumerina (anamorph Plectosporium tabacinum) was isolated from a urine sample. Clinical manifestations were blindness, altered mentation, and osteomyelitis. Treatment with itraconazole and terbinafine for greater than one year resulted in stable clinical disease.

Rasamsonia argillacea pulmonary and aortic graft infection in an immune-competent patient.

Rasamsonia argillacea (formerly known as Geosmithia argillacea) is a fungus recently recognized as a pathogen of immunocompromised patients. Here we report the first case of Rasamsonia infection in an immunocompetent host, presenting as a pulmonary and aortic graft infection. Its morphological similarity to nonpathogenic Penicillium species delayed the diagnosis and initiation of appropriate treatment.

The Changing Epidemiology of Oropharyngeal Candidiasis in Patients with HIV/AIDS in the Era of Antiretroviral Therapy.

The impact of antiretroviral therapy (ART) on opportunistic conditions in HIV patients continues to evolve. We specifically studied the changing epidemiology of oropharyngeal candidiasis (OPC) in 215 HIV/AIDS patients. Status of yeast colonization was assessed from oral rinse samples, and preliminary yeast identification was made using CHROMagar Candida and confirmed with standard microbiological techniques and/or molecular sequencing. Susceptibility to fluconazole was determined by CHROMagar Candida agar dilution screening and CLSI broth microdilution. 176 (82%) patients were colonized and 59 (27%) patients had symptomatic OPC. Candida albicans was the most prevalent species, though C. glabrata and C. dubliniensis were detected in 29% of isolates. Decreased fluconazole susceptibility occurred in 10% of isolates. Previous ART reduced the risk of OPC, while smoking increased the risk of colonization. Oral yeast colonization and symptomatic infection remain common even with advances in HIV therapy. C. albicans is the most common species, but other yeasts are prevalent and may have decreased susceptibility to fluconazole.

Invasive Mycoleptodiscus fungal cellulitis and myositis.

We report progressive necrotizing fungal cellulitis and myositis in the leg of a patient with glioblastoma multiforme treated with temozolomide and corticosteroids. While the morphologic appearance of the isolate and its ability to grow at temperatures greater than 32°C were suggestive of Mycoleptodiscus indicus, some of the conidia were atypical for this species in that they had single septa and occasional lateral appendages. Furthermore, the isolate was different from M. indicus based on the sequencing analysis of two rDNA regions. This is the first case of Mycoleptodiscus invasive fungal disease in which the causative agent could not be resolved at the species level because of inconsistencies between morphological and molecular data.

Graphium basitruncatum fungemia in an immunosuppressed child post stem-cell transplantation.

Graphium basitruncatum is genetically and morphologically distinct from other Graphium and Scedosporium species, and has been reported only once previously as a cause of human disease. We report a case of Graphium basitruncatum fungemia in a two year old child with dyskeratosis congenita who underwent stem cell transplantation two months prior to infection.

Geosmithia argillacea: an emerging cause of invasive mycosis in human chronic granulomatous disease.

BACKGROUND: Chronic granulomatous disease (CGD) is an inherited disorder of the nicotinamide adenine dinucleotide phosphate oxidase that leads to defective production of microbicidal superoxide and other oxidative radicals, resulting in increased susceptibility to invasive infections, especially those due to fungi. METHODS: Geosmithia argillacea was identified from cultured isolates by genomic sequencing of the internal transcribed spacer region. Isolates previously identified as Paecilomyces variotii, a filamentous fungus closely resembling G. argillacea, were also examined. RESULTS: We identified G. argillacea as the cause of invasive mycosis in 7 CGD patients. In 5 cases, the fungus had been previously identified morphologically as P. variotii. All patients had pulmonary lesions; 1 had disseminated lesions following inhalational pneumonia. Infections involved the chest wall and contiguous ribs in 2 patients and disseminated to the brain in 1 patient. Four patients with pneumonia underwent surgical intervention. All patients responded poorly to medical treatment, and 3 died. CONCLUSIONS: We report the first cases of invasive mycosis caused by G. argillacea in CGD patients. G. argillacea infections in CGD are often refractory and severe with a high fatality rate. Surgical intervention has been effective in some cases. G. argillacea is a previously underappreciated and frequently misidentified pathogen in CGD that should be excluded when P. variotii is identified morphologically.

A case of bovine valve endocarditis caused by Engyodontium album.

We report the first case of Engyodontium album bioprosthetic valve endocarditis in a 44-year-old male with a history of juvenile rheumatoid arthritis. There is only one other report of Engyodontium album as a human pathogen. The present case supports the increased incidence of fungal endocarditis especially in patients receiving immunotherapy.

Disseminated human conidiobolomycosis due to Conidiobolus lamprauges.

We describe a disseminated fungal infection by Conidiobolus lamprauges in a patient with malignant lymphoma. Histopathology and mycological studies were performed, along with molecular analyses. This is the first record of this species causing human disease and the fifth reported disseminated infection by a Conidiobolus sp. in humans.

Cerebral phaeohyphomycosis due to Rhinocladiella mackenziei (formerly Ramichloridium mackenziei): a taxonomic update and review of the literature.

Cerebral phaeohyphomycosis caused by Rhinocladiella mackenziei (formerly Ramichlo-ridium mackenziei) is extremely rare, and geographically limited to the Middle East. The fungus exclusively targets the brain and infections have a grave prognosis. Eighteen cases have been reported in the literature from 1983 to 2004 with almost 100% mortality. Our patient presented in February 2008 with a brain abscess while receiving chemotherapy for carcinoma of the breast. Diagnosis was by craniotomy and aspiration of the brain abscess. Direct microscopy showed dematiaceous fungal hyphae. R. mackenziei was recovered in culture and this identification was confirmed by molecular analysis. Examination of histopathological sections of tissue from the brain biopsy revealed moniliform hyphae characteristic for phaeohyphomycosis. The patient failed to respond to antifungal therapy with amphotericin B and voriconazole or amphotericin B and posaconazole and finally expired 64 days after diagnosis. In vitro antifungal susceptibility testing showed this isolate to be resistant to amphotericin B while susceptible to itraconazole, voriconazole, and posaconazole. Previously published antifungal susceptibility data indicate that although strains show variable susceptibility to amphotericin B, the organism is generally refractory to treatment with this agent. Similar outcomes are seen with the azole agents used alone or in combination with other drugs. Although no specific risk factors have been identified, the majority of cases have occurred in immunocompromised individuals. R. mackenziei is a highly virulent agent of serious cerebral phaeohyphomycosis, and should be considered in the differential diagnosis of central nervous system disease in the Middle East.

Beauveria keratitis and biopesticides: case histories and a random amplification of polymorphic DNA comparison.

PURPOSE: The purposes of this study were to describe 2 contact lens-associated Beauveria keratitis cases and to compare the isolates of 3 contact lens-associated Beauveria keratitis cases with Beauveria-based biopesticides using random amplification of polymorphic DNA (RAPD). METHODS: A 55-year-old diabetic woman from New Mexico and a 31-year-old healthy woman from southern Wisconsin developed soft contact lens-related corneal ulcers unresponsive to topical moxifloxacin and prednisolone acetate drops. Their corneal cultures grew B. bassiana. These isolates, an isolate from a third soft contact lens-related Beauveria keratitis case, and Beauveria-based biopesticides sold in the United States were analyzed using morphological features, DNA sequencing, and RAPD. A PubMed, Cochrane Library, OVID, UpToDate, and Google search using the term "Beauveria" found only 9 reported Beauveria keratitis infections. RESULTS: Patient 1 responded to topical natamycin, ketoconazole, and 200 mg oral ketoconazole twice daily before developing a secondary bacterial infection requiring penetrating keratoplasty. After subsequent cataract surgery, the best-corrected visual acuity was 20/20. Patient 2 was treated with topical natamycin, topical amphotericin, and 200 mg oral voriconazole twice daily for 1 month with residual scarring and a best-corrected visual acuity of 20/25. RAPD showed that all isolates were unrelated. CONCLUSIONS: Although earlier reported Beauveria keratitis cases occurred after corneal injury in patients who did not wear contact lenses, 3 recent patients wore soft contact lenses and denied trauma, mirroring a changing trend in microbial keratitis. RAPD analysis showed that the Beauveria isolates were unrelated to one another and to Beauveria-based biopesticides. In Patient 2, oral voriconazole worked well.