RESUMEN
BACKGROUND: Group B Streptococcus (GBS) infections caused by Streptococcus agalactiae is a leading cause of meningitis and sepsis in neonates, with early-onset GBS symptoms emerging during the first week of life and late-onset occurring thereafter. Perinatal transmission of GBS to the neonate through the birth canal is the main factor associated with early-onset neonate infections, while less is understood about the source of late-onset infections. METHODS: In this report we describe a case of twin ex-premature infants who presented one month after birth with GBS septicemia. The mother had been appropriately screened at gestational age 35-37 weeks and laboratory methods failed to detect GBS colonization by culture or clinical molecular methods. In attempts to identify and isolate the source of GBS infection, additional surveillance swabs were collected from the mother at the time of neonate admission. Culture and a commercially available, FDA-cleared molecular PCR assay were performed. RESULTS: No GBS was detected from swabs collected from the perianal, thigh/groin or axillary areas. However, expressed breast milk and swabs from the breastmilk pump were positive by both methods. Since simultaneous culture and molecular methods which used breastmilk as a source were performed, investigators ascertained the limit of detection for GBS in breastmilk. The limit of detection was determined to be tenfold lower than that of LIM-broth enriched cultures-the FDA-approved source. Subsequent whole genome sequencing (WGS) analysis of isolates recovered from breastmilk and blood cultures from the infants demonstrated all strains were related and characterized as ST-452. Both infants responded very well to treatment and continued to have no related events or concerns at the two-year follow up appointment. CONCLUSIONS: Strain type 452 (capsular type IV) has recently emerged as a hypervirulent strain and has previously been documented as causing GBS infections in elderly populations. Antibiotic therapy resolved both mother and infant infections. Subsequent testing for the presence of GBS in breastmilk samples also showed an absence of bacteria. This is the first report of infant twins late-onset GBS infections caused by the hypervirulent S. agalactiae ST-452 with breastmilk as the source.
Asunto(s)
Bacteriemia/microbiología , Enfermedades del Recién Nacido/microbiología , Leche Humana/microbiología , Infecciones Estreptocócicas/transmisión , Streptococcus agalactiae/genética , Streptococcus agalactiae/aislamiento & purificación , Adulto , Bacteriemia/sangre , Bacteriemia/diagnóstico , Bacteriemia/transmisión , Sangre/microbiología , Extracción de Leche Materna , Femenino , Humanos , Recién Nacido , Enfermedades del Recién Nacido/sangre , Enfermedades del Recién Nacido/diagnóstico , Recien Nacido Prematuro/sangre , Transmisión Vertical de Enfermedad Infecciosa , Masculino , Técnicas de Diagnóstico Molecular , Filogenia , Infecciones Estreptocócicas/sangre , Infecciones Estreptocócicas/diagnóstico , Infecciones Estreptocócicas/microbiología , Streptococcus agalactiae/clasificación , Streptococcus agalactiae/patogenicidad , VirulenciaRESUMEN
In 2017, a mumps outbreak occurred in a barrack holding 249 service members. Suspected cases were evaluated with a combination of mumps IgG, IgM, viral culture, PCR and sequencing. Seven cases were diagnosed in febrile patients presenting with parotitis or orchitis. Mumps infection was confirmed by IgM or positive PCR with 5/7 cases having notable IgG levels before infection. Sequencing confirmed mumps genotype G strain. Serum from all 249 service members collected prior to the outbreak was withdrawn from the Department of Defense (DoD) Serum Repository and the IgG values of measles, mumps and rubella determined with 20.2%, 12.3% and 9.7% service members being seronegative, respectively. No specific IgG seronegativity combination predicted IgG marker levels to another virus within the same vaccine. This paper provides additional evidence that mumps serology is not a reliable surrogate for mumps immunity and that we need better laboratory correlates to confirm immunity.
Asunto(s)
Anticuerpos Antivirales/sangre , Vacuna contra el Sarampión-Parotiditis-Rubéola/inmunología , Virus de la Parotiditis/inmunología , Paperas/inmunología , Adulto , Brotes de Enfermedades , Femenino , Humanos , Inmunoglobulina G/sangre , Inmunoglobulina M/sangre , Masculino , Personal Militar , Morbillivirus/inmunología , Paperas/epidemiología , Virus de la Parotiditis/genética , Virus de la Rubéola/inmunología , Vacunación , Adulto JovenRESUMEN
Clostridium difficile can carry a genetically variable pathogenicity locus (PaLoc), which encodes clostridial toxins A and B. In hospitals and in the community at large, this organism is increasingly identified as a pathogen. To develop a diagnostic test that combines the strengths of immunoassays (cost) and DNA amplification assays (sensitivity/specificity), we targeted a genetically stable PaLoc region, amplifying tcdB sequences and detecting them by hybridization capture. The assay employs a hot-start isothermal method coupled to a multiplexed chip-based readout, creating a manual assay that detects toxigenic C. difficile with high sensitivity and specificity within 1 h. Assay automation on an electromechanical instrument produced an analytical sensitivity of 10 CFU (95% probability of detection) of C. difficile in fecal samples, along with discrimination against other enteric bacteria. To verify automated assay function, 130 patient samples were tested: 31/32 positive samples (97% sensitive; 95% confidence interval [CI], 82 to 99%) and 98/98 negative samples (100% specific; 95% CI, 95 to 100%) were scored correctly. Large-scale clinical studies are now planned to determine clinical sensitivity and specificity.
Asunto(s)
Automatización de Laboratorios/métodos , Proteínas Bacterianas/genética , Toxinas Bacterianas/genética , Técnicas Bacteriológicas/métodos , Clostridioides difficile/aislamiento & purificación , Análisis por Micromatrices/métodos , Técnicas de Diagnóstico Molecular/métodos , Técnicas de Amplificación de Ácido Nucleico/métodos , Clostridioides difficile/genética , Infecciones por Clostridium/diagnóstico , Infecciones por Clostridium/microbiología , Heces/microbiología , Humanos , Sensibilidad y Especificidad , Factores de TiempoRESUMEN
We present the evolution of testing algorithms at our institution in which the C. Diff Quik Chek Complete immunochromatographic cartridge assay determines the presence of both glutamate dehydrogenase and Clostridium difficile toxins A and B as a primary screen for C. difficile infection and indeterminate results (glutamate dehydrogenase positive, toxin A and B negative) are confirmed by the GeneXpert C. difficile PCR assay. This two-step algorithm is a cost-effective method for highly sensitive detection of toxigenic C. difficile.
Asunto(s)
Técnicas Bacteriológicas/métodos , Clostridioides difficile/aislamiento & purificación , Infecciones por Clostridium/diagnóstico , Infecciones por Clostridium/microbiología , Algoritmos , Proteínas Bacterianas/genética , Proteínas Bacterianas/inmunología , Toxinas Bacterianas/genética , Toxinas Bacterianas/inmunología , Enterotoxinas/genética , Enterotoxinas/inmunología , Glutamato Deshidrogenasa/genética , Glutamato Deshidrogenasa/inmunología , Hospitales Universitarios , Humanos , Inmunoensayo/métodos , Reacción en Cadena de la Polimerasa/métodosRESUMEN
The Cepheid Xpert MTB/RIF research-use-only (RUO) assay and a laboratory-developed test (LDT) targeting IS6110 were evaluated and compared to mycobacterial culture as the gold standard. The performance characteristics of both molecular assays were determined by using 112 specimens from 90 patients, including 89 pulmonary specimens and 23 extrapulmonary specimens. Of the specimens tested, 37 (33%) were culture positive for the Mycobacterium tuberculosis complex; 29 were pulmonary, and 8 were extrapulmonary. Of these culture-positive specimens, 83% of the pulmonary specimens and 50% of the extrapulmonary specimens were smear positive. There was complete concordance between the smear-positive culture-positive specimens, independent of the anatomical site (100% sensitivity). The sensitivity of the MTB/RIF RUO assay for smear-negative specimens was 60% for pulmonary and 75% for extrapulmonary specimens, while the IS6110 LDT sensitivities were 40% and 0%, respectively. There was also complete concordance among the culture-negative specimens tested. Both assays showed 95% specificity, with four culture-negative specimens testing as positive. A review of patient records indicated that there was a high likelihood of the presence of M. tuberculosis complex DNA in the false-positive specimens. Biosafety analysis was performed and showed an acceptable reduction in organism viability using the processing methods described above. Both molecular assays are suitable for the detection of M. tuberculosis isolates in smear-positive pulmonary and extrapulmonary specimens, while the sensitivity of the detection of M. tuberculosis isolates in smear-negative specimens was variable.
Asunto(s)
Técnicas Bacteriológicas/métodos , Técnicas de Diagnóstico Molecular/métodos , Mycobacterium tuberculosis/aislamiento & purificación , Reacción en Cadena en Tiempo Real de la Polimerasa/métodos , Tuberculosis/diagnóstico , Elementos Transponibles de ADN , ADN Bacteriano/genética , Humanos , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/crecimiento & desarrollo , Sensibilidad y Especificidad , Tuberculosis/microbiologíaRESUMEN
In this work, we report the cloning and characterization of the first cell surface casein kinase II (CKII) substrate (Tc-1) of Trypanosoma cruzi, the causative agent of Chagas' disease. Analysis of the gene sequence revealed a 1,653-bp open reading frame coding for 550 amino acid residues. Northern blot analysis showed a 4.5-kb transcript that is expressed in invasive trypomastigotes but not in noninvasive epimastigote forms of T. cruzi. Southern blot analysis indicates that Tc-1 is a single-copy gene. At the amino acid level, Tc-1 displayed 95% and 99% identity to two hypothetical proteins recently reported by the T. cruzi genome project. Analysis of the translated amino acid sequence indicates that the Tc-1 gene has a putative transmembrane domain with multiple cytoplasmic and extracellular CKII phosphosites. Exogenous human CKII was able to phosphorylate serine residues on both recombinant Tc-1 and Tc-1 of intact trypomastigotes. This phosphorylation was inhibited by the CKII inhibitors heparin and 4,5,6,7,-tetrabromo-2-azabenzimidazole. Immunoblots of solubilized trypomastigotes, epimastigotes, and amastigotes probed with anti-recombinant Tc-1 immunoglobulin G revealed a 62-kDa protein that is expressed only in infective trypomastigotes. Immunoprecipitation of labeled surface proteins of trypomastigotes indicated that the 62-kDa protein is a surface protein, and we found that the protein is uniformly distributed on the surface of trypomastigotes by direct immunofluorescence. Antibodies to Tc-1 effectively blocked trypomastigote invasion of host cells and consequently reduced parasite load. Preincubation of either trypomastigotes or myoblasts with CKII inhibitors blocked T. cruzi infection. Thus, for the first time, we describe a cell surface CKII substrate of a protozoan parasite that is phosphorylated by human CKII and that is involved in cellular infection.
Asunto(s)
Quinasa de la Caseína II/metabolismo , Clonación Molecular , Proteínas de la Membrana/genética , Mioblastos Cardíacos/parasitología , Proteínas Protozoarias/genética , Trypanosoma cruzi/genética , Trypanosoma cruzi/metabolismo , Secuencia de Aminoácidos , Animales , Quinasa de la Caseína II/química , Quinasa de la Caseína II/genética , Quinasa de la Caseína II/aislamiento & purificación , Humanos , Proteínas de la Membrana/aislamiento & purificación , Proteínas de la Membrana/metabolismo , Datos de Secuencia Molecular , Peso Molecular , Mioblastos Cardíacos/enzimología , Fosforilación , Proteínas Protozoarias/aislamiento & purificación , Proteínas Protozoarias/metabolismo , Ratas , Especificidad por Sustrato , Trypanosoma cruzi/crecimiento & desarrollo , Trypanosoma cruzi/patogenicidad , Factores de Virulencia/genética , Factores de Virulencia/aislamiento & purificaciónRESUMEN
OBJECTIVE: Distal ischemic necrosis of the flap remains an unsolved, challenging problem. Phosphodiesterase (PDE) inhibitors, which include the drug sildenafil, are a relatively new class of U.S. Food and Drug Administration-approved medications whose effect on tissue viability has not been widely explored. The vasodilatory effects of these drugs have the potential to enhance blood flow to flaps and increase their survivability. The purpose of this study was to examine the short- and long-term effects of sildenafil, administered intraperitoneally at a dose of 9 mg/kg per day, on the survival of surgical skin flaps in rats. METHODS: A McFarlane-type random pattern skin (3 x 10-cm) flap model was used to evaluate the effect of sildenafil on necrosis at multiple time points. Rats were assigned to sildenafil-treated (9 mg/kg per day intraperitoneally; n = 34), vehicle control (n = 35), or sham (no injection; n = 40) groups. In each group, caudally based, dorsal, rectangular (3 x 10-cm) flaps were created. Flap necrosis was determined using orthogonal polarization spectral imaging and digital photography analysis on days 1, 3, 5, and 7 postsurgery. RESULTS: Orthogonal polarization spectral imaging results showed a significant decrease in necrosis and stasis in rats treated with sildenafil on days 1 and 3. Although reductions observed at days 5 and 7 were not as dramatic as days 1 and 3, digital photography analysis confirmed a decrease in the area of necrosis at all time points evaluated. CONCLUSIONS: These results suggest that PDE 5 inhibitors may play a more important role in early postoperative skin flap viability rather than at later time points and may be beneficial for skin flap viability as shown in the rat model. PDE 5 inhibitors may reduce the extent of necrosis after reconstructive surgeries.
Asunto(s)
Procedimientos Quirúrgicos Dermatologicos , Microcirculación/efectos de los fármacos , Inhibidores de Fosfodiesterasa/farmacología , Piperazinas/farmacología , Colgajos Quirúrgicos/irrigación sanguínea , 3',5'-GMP Cíclico Fosfodiesterasas/antagonistas & inhibidores , Animales , Velocidad del Flujo Sanguíneo/efectos de los fármacos , Modelos Animales de Enfermedad , Estudios de Seguimiento , Masculino , Necrosis/patología , Necrosis/prevención & control , Purinas , Ratas , Ratas Sprague-Dawley , Citrato de Sildenafil , Sulfonas , Colgajos Quirúrgicos/patología , Factores de Tiempo , Resultado del TratamientoRESUMEN
BACKGROUND: Concern about accidental contact transmission after smallpox vaccination has prompted various recommendations regarding vaccination site coverage. METHODS: On days 6-8 after their first-ever smallpox vaccination, 63 adult subjects were randomized to apply a self-adhesive bandage (n=20), gauze with adhesive tape (n=21), or gauze with a semipermeable dressing (n=22) over the vaccination site for a mean of 8+/-2 h. Swabs from the external bandage surfaces and the vaccination sites were then assessed by real time vaccinia-specific polymerase chain reaction (PCR) in blinded fashion. RESULTS: Among 58 subjects completing the study, PCR results were positive for the vaccination site in 55 (94.8%) and on 10 swabs (17.2%) from external bandage surfaces. There were no differences among the 3 bandages (P=.57). CONCLUSIONS: At 7 days after smallpox vaccination, a peak time for vaccinia shedding, a self-adhesive bandage was as effective as 2 bulkier, less convenient bandages in limiting PCR-detectable virus on the external surface.
Asunto(s)
Vendajes/virología , Vacuna contra Viruela/administración & dosificación , Vacunación/métodos , Virus Vaccinia/aislamiento & purificación , Vaccinia/prevención & control , Vaccinia/transmisión , Adulto , ADN Viral/aislamiento & purificación , Femenino , Humanos , MasculinoRESUMEN
Host growth factors induce proliferation of Trypanosoma cruzi amastigotes by mechanisms that remain poorly defined. Here we examined human epidermal growth factor (EGF) for its ability to bind to the mammalian multiplicative forms of T. cruzi and to induce growth of the parasites. EGF stimulated incorporation of [3H] thymidine into DNA and growth of amastigotes both in a concentration-dependent manner. Radiolabeled EGF was found to bind to amastigotes in a concentration-dependent and saturable manner but it did not bind to trypomastigotes. Scatchard analysis showed a single class of receptors with a Kd of 0.8 nM and numbering 3.1 x 10(3) per amastigote. Results from internalization experiments provided evidence of receptor-mediated endocytosis of EGF. Northern analysis showed a 3.0-kb transcript for the putative EGF receptor (EGFR) homologue in amastigotes, but not trypomastigotes. Binding of EGF to amastigotes induced signal transduction events. EGF induced "in vitro" kinase activity as determined by gamma-[32P] ATP incorporation into amastigote proteins. EGF also increased protein kinase C activity in a concentration-dependent manner and Mitogen Activated Protein (MAP) kinase activity in a time- and concentration-dependent manner. A specific inhibitor (AG14782) of the EGFR and a MAP kinase inhibitor (PD98059) decreased EGF-dependent T. cruzi MAP kinase activity. These results describe a novel mechanism used by amastigotes to regulate their proliferation mediated by an EGF-dependent signal transduction pathway.