Efficacy of disinfection procedures to reduce Acinetobacter baumanii blaOXA-23 contamination rate of needleless connectors: an in-vitro study.

Aim: This study aimed to verify the efficacy of disinfection procedures to reduce Acinetobacter baumannii blaOXA-23 bacterial load in needleless connectors that had been experimentally contaminated. Methods: Two-way intermediate extender's hub and needle-free valve were contaminated with Acinetobacter baumannii blaOXA-23. To disinfect them, the following procedures were carried out: sterile gauze with 70% ethanol, sterile gauze with Incidin®, and 70% isopropyl alcohol single-use cap, with eight times friction for 10 s, followed by 5 s drying time. The statistical tests Kruskal-Wallis and post-hoc Conover were performed using MedCalc®. Results: A total of 82 experiments were conducted. All tested disinfection procedures were efficacious in reducing the A. baumannii blaOXA-23 load. The 70% IPA single-use cap was found to be the best method for disinfecting the two-way intermediate extender's hub (87.28%), while all the methods were efficacious for the disinfection of the needle-free valve (more than 90%). During the inoculation period, A. baumannii blaOXA-23 showed less adherence to the needle-free valve during the inoculation period, probably due to the device's design. Conclusion: The three tested disinfection procedures using sterile gauze with 70% ethanol, sterile gauze with Incidin®, and 70% IPA single-use cap were found to be efficacious in reducing the bacterial load of A. baumanni blaOXA-23 in needleless connectors. Proper disinfection of needleless connectors is a crucial nursing practice to prevent bloodstream infections, as it significantly reduces the bacterial load present in the device.

Impact of mutations in hVISA isolates on decreased susceptibility to vancomycin, through population analyses profile - area under curve (PAP-AUC).

We analyzed sequences of graSR, vraSR, walKR and rpoB genes in hVISA from Brazil. Five isolates showed mutations in at least one gene. rpoB H481N and graS T224I were the most frequent mutations, followed by graR D148Q and walK A468T. Our study reinforces the heterogeneity of genetic patterns among hVISA.

PCR-RFLP assay as an option for primary HPV test.

Persistent human papillomavirus (HPV) infection is an essential factor of cervical cancer. This study evaluated the analytical performance of restriction fragment length polymorphism polymerase chain reaction (PCR-RFLP) assay compared to PapilloCheck® microarray to identify human papilloma virus (HPV) in cervical cells. Three hundred and twenty-five women were analyzed. One sample was used for conventional cytology and another sample was collected using BD SurePath™ kit for HPV tests. Eighty samples (24.6%) were positive for HPV gene by PCR-Multiplex and were then submitted to PCR-RFLP and PapilloCheck® microarray. There was a genotyping agreement in 71.25% (57/80) on at least one HPV type between PCR-RFLP and PapilloCheck® microarray. In 22 samples (27.5%), the results were discordant and those samples were additionally analyzed by DNA sequencing. HPV 16 was the most prevalent HPV type found in both methods, followed by HPVs 53, 68, 18, 39, and 66 using PCR-RFLP analysis, and HPVs 39, 53, 68, 56, 31, and 66 using PapilloCheck® microarray. In the present study, a perfect agreement using Cohen's kappa (κ) was found in HPV 33 and 58 (κ=1), very good for HPV 51, and good for types 16, 18, 53, 59, 66, 68, 70, and 73. PCR-RFLP analysis identified only 25% (20/80) HPV coinfection, and PapilloCheck® microarray found 62.5% (50/80). Our Cohen's kappa results indicate that our in-house HPV genotyping testing (PCR-RFLP analysis) could be applied as a primary HPV test screening, especially in low income countries. If multiple HPV types are found in this primary test, a more descriptive test, such as PapilloCheck® microarray, could be performed.

Detection of hepatitis A virus (HAV) in oysters (Crassostrea gigas).

Because shellfish (oysters, clams, and mussels) are filter-feeders, pathogens become concentrated within them, and human consumption of raw, or under-cooked shellfish can result in disease outbreaks. Identification of hepatitis A virus (HAV) in shellfish has been difficult for several reasons: the concentration of virions in shellfish tissues are very low, detection methods based on in vitro propagation are unreliable, recovery of virions from shellfish tissues is inefficient, and PCR inhibitors in shellfish tissues limit the success of RT-PCR. These facts underlie difficulties in determining cause and effect relationships between hepatitis A outbreaks and detection of HAV contamination in shellfish samples. We have developed a reliable and highly sensitive method for detection of HAV in oyster tissues at low levels (0.001 FFU/ml-fluorescent focus units per milliliter). Our method combines dissection of the gastrointestinal oyster tract, organic extraction before PEG precipitation, and RNA extraction with Trizol LS, followed by RT-PCR and hybridization using a digoxigenin-labeled HAV cDNA probe. Our results will benefit both public health officials concerned about hepatitis A infections caused by consumption of HAV-contaminated oysters and shellfish producers who require reliable methods for quality control of commercial oyster production.

Antiviral evaluation of plants from Brazilian Atlantic Tropical Forest.

The antiviral activity of six medicinal plants from Brazilian Atlantic Tropical Forest was investigated against two viruses: herpes simplex virus type 1 (HSV-1) and poliovirus type 2 (PV-2). Cuphea carthagenensis and Tillandsia usneoides extracts showed the best antiherpes activity. T. usneoides dichloromethane, ethyl acetate and n-butanol extracts, and Lippia alba n-butanol extract showed inhibition of HSV-1, strain 29R/acyclovir resistant. In addition, only L. alba ethyl acetate extract showed antipoliovirus activity. These results corroborate that medicinal plants can be a rich source of potential antiviral compounds.

Detection of adenoviruses in shellfish by means of conventional-PCR, nested-PCR, and integrated cell culture PCR (ICC/PCR).

We tested three PCR based methodologies to detect adenoviruses associated with cultivated oysters. Conventional-PCR, nested-PCR, and integrated cell culture-PCR (ICC/PCR) were first optimized using oysters seeded with know amounts of Adenovirus serotype 5 (Ad5). The maximum sensitivity for Ad5 detection was determined for each method, and then used to detect natural adenovirus contamination in oysters from three aquiculture farms in Florianopolis, Santa Catarina State, Brazil, over a period of 6 months. The results showed that the nested-PCR was more sensitive (limit of detection: 1.2 PFU/g of tissue) than conventional-PCR and ICC-PCR (limit of detection for both: 1.2 x 10(2)PFU/g of tissue) for detection of Ad5 in oyster extracts. Nested-PCR was able to detect 90% of Ad5 contamination in harvested oyster samples, while conventional-PCR was unable to detect Ad5 in any of the samples. The present work suggests that detection of human adenoviruses can be used as a tool to monitor the presence of human viruses in marine environments where shellfish grow, and that nested-PCR is the method of choice.