Short communication: High incubation temperature in bovine mammary epithelial cells reduced the activity of the mTOR signaling pathway.

Hyperthermia alters utilization of AA in protein synthesis and cell-signaling activity in bovine mammary cells. Essential AA and insulin regulate translation of proteins by controlling the activity of mammalian target of rapamycin (mTOR) signaling pathway. The objectives of this study were to evaluate (1) the effects of incubation temperature on the mTOR signaling pathway and transcription of AA transporters in a bovine mammary alveolar cell line (MAC-T) and (2) the combined effects of incubation temperature and insulin on the mTOR signaling pathway in this cell line. Cells were cultured in medium with 10% fetal bovine serum at 37°C and 5% CO2. In experiment 1, cells were subjected to 37°C (control) or 41.5°C (high incubation temperature; HT) for 12 h. In experiment 2, cells were assigned to 1 of 4 treatments as a 2 × 2 factorial arrangement, including 2 cell culture temperatures (control and HT) and absence or presence of 1.0 µg/mL of insulin. Proteins were harvested and separated by gel electrophoresis. In experiment 1, gene expression of AA transporters (SLC1A1, SLC1A5, SLC3A2, SLC7A1, SLC7A5, and SLC36A1) were evaluated, and changes of ≥2 fold were deemed significantly different. In experiments 1 and 2, immunoblotting was used to identify total and site-specific phosphorylated forms of protein kinase B (Akt1; Ser473), p70 S6 kinase (S6K1; Thr389), ribosomal protein S6 (rpS6; Ser235/236), and eukaryotic elongation factor 2 (eEF2; Thr56). Phosphorylated and total forms of Akt1, S6K1, rpS6, and eEF2 were quantified and expressed as the ratio of phosphorylated to total protein. In experiment 1, HT resulted in a ≥2-fold increase expression of SLC1A1 and SLC3A2. High incubation temperature reduced the phosphorylated to total ratio of Akt1 and rpS6 and increased the phosphorylated to total ratio of eEF2. In experiment 2, we found no temperature by insulin interactions on phosphorylation state of the protein factors of interest. High incubation temperature reduced the phosphorylated to total ratio of Akt1. The addition of insulin increased the phosphorylated to total ratio of Akt1, S6K1, and rpS6. In summary, HT reduced the activity of the mTOR signaling pathway and increased the expression of AA transporters. High incubation temperature possibly reduced protein translation by reducing the mTOR signaling pathway activity in an effort to adapt to thermal stress. These results may help explain the direct effect of elevated temperature on AA metabolism and protein translation in heat-stressed animals.

Bovine intramammary infection associated immunogenic surface proteins of Streptococcus uberis.

In spite of the increasing prevalence of Streptococcus uberis mastitis, its pathogenesis and associated virulence factors are not clearly defined. The aim of this study was to identify virulence associated genes and their products that can be used to develop effective vaccine to control bovine S. uberis mastitis. S. uberis was co-cultured with primary bovine mammary epithelial cells (PBMEC) or infused into mammary gland of dairy cows. The messenger RNA (mRNA) from S. uberis associated with PBMEC after 2 h or 4 h of co-culture was purified and sequenced. Results showed that virulence-associated genes such as surface lipoprotein (slp), infection induced histidine kinase (iihK), infection induced response regulator (iirR) and extracellular sugar binding protein 1 and 2 (exsbP1 and exsbP2) were among the top-up-regulated genes. To verify this observation in vivo, quantitative real time PCR (qRT - PCR) was conducted on mRNA of S. uberis recovered from milk of infected mammary glands 24 h post infection. Results revealed that in vitro up-regulated virulence-associated genes were also significantly up regulated under in vivo conditions. The iihK and iirR are flanked by exsbP1 and exsbP2 genes and this entire operon seems to be involved in adaptation to glands micro-environment, survival and colonization of the bovine mammary glands. Based on immunogenic epitope prediction of proteins encoded by these up-regulated genes during early stages of host-bacterial interactions slp, exsbP1 and exsbP2 genes were selected, cloned and expressed in E. coli. The purified recombinant proteins (rSlP, rExsbP1 & rExsbP2) reacted strongly with convalescent serum from cows experimentally infected with S. uberis confirming that they are immunogenic. These proteins may serve as potential targets to develop an effective vaccine against S. uberis mastitis.

A case report on management of synergistic gangrene following an incisional abdominal hernia repair in an immunocompromised obese patient.

INTRODUCTION: We present a case on conservative management of salvaging the mesh in an immunocompromised morbidly obese patient, who developed a synergistic gangrene infection following a primary open mesh repair of an incisional hernia. PRESENTATION OF CASE: Our patient presented with a surgical wound infection, comorbidities were Chronic Lymphoblastic Leukemia (CLL), Body Mass Index (BMI) of 50, hypertension and diet controlled type-2 diabetes. In surgery, wide necrotic wound debridement, early and repetitive wound drainages with the use of a large pore polypropylene mesh and a detailed surgical follow up was required. High dose intravenous broad-spectrum antibiotic treatment and Negative Pressure Wound Therapy (NPWT) was administrated in combination with adopting a multidisciplinary approach was key to our success. DISCUSSION: Stoppa Re et al. complied a series of 360 ventral hernia mesh repairs reporting an infection rate of 12% that were managed conservatively. However, our selective case is unique within current literature, being the first to illustrate mesh salvage in a morbid obese patient with CLL. Recent modifications in mesh morphology, such as lower density, wide pores, and lighter weight has led to considerable improvements regarding infection avoidance. CONCLUSION: This case has demonstrated how a planned multidisciplinary action can produce prosperous results in a severely obese immunocompromised patient with an SSI, following an incisional hernia repair.

Short communication: Evaluation of bulk tank milk microbiological quality of nine dairy farms in Tennessee.

The purpose of this study was to evaluate the bulk tank milk (BTM) quality of 9 East Tennessee dairy farms and to determine its relationship with selected quality milk parameters. Bulk tank milk samples (n=1,141) were collected over a 42-mo period (June 2006 through November 2009) from farms, based on their preliminary incubation count (PIC) history. Parameters of BTM quality evaluated in this study included somatic cell count (SCC), standard plate count (SPC), PIC, laboratory pasteurization count (LPC), Staphylococcus spp. count, Streptococcus spp. count, and coliform count. Strong correlations between SPC and Streptococcus spp. counts (0.72) and between SPC and PIC (0.70) were found. However, moderate correlations were seen among other milk quality parameters. In addition, seasonal variations for some milk quality parameters were noted. For example, milk quality parameters such as SCC, SPC, LPC, and coliform count were significantly higher in summer, whereas Streptococcus spp. counts were significantly higher in winter. No seasonal variation in PIC or Staphylococcus spp. counts was observed. Summarizing, results from this investigation showed the importance of using several bacterial counts (SCC, SPC, PIC, LPC, Streptococcus spp. count, Staphylococcus spp. count, and coliform counts) as simultaneous indicators of milk quality.

Historical perspective and human consequences of Africanized bee stings in the Americas.

In 1956, Africanized bees began to spread in the American continent from southern Brazil, where original African bees mated with European bees. A few years later, in 1990, these Africanized bees reached the United States and were found in Texas. Currently, these hybrid bees are found in several North American states and will probably reach the Canadian border in the future. Although the presence of Africanized bees had produced positive effects on Brazilian economy, including improvement in crop pollination and in honey production, turning Brazil into a major exporter, the negative impacts-such as swarming, aggressive behavior, and the ability to mass attack-resulted in serious and fatal envenomation with humans and animals. Victims of bee attacks usually develop a severe envenomation syndrome characterized by the release of a large amount of cytokines [interleukins (IL) IL-1, IL-6, IL-8], and tumor necrosis factor (TNF). Subsequently, such cytokines produce an acute inflammatory response that triggers adverse effects on skeletal muscles; bone marrow; hepatic and renal functions; and cardiovascular, central nervous, and immune systems. Finally, the aim of the present review is to study historical characteristics and current status of Africanized bees' spread, the composition of their venom, the impact of the bees on the Brazilian economy and ecology, and clinical aspects of their stings including immune response, and to suggest a protocol for bee sting management since there is no safe and effective antivenom available.

Host-pathogen gene expression profiles during infection of primary bovine mammary epithelial cells with Escherichia coli strains associated with acute or persistent bovine mastitis.

Escherichia coli intramammary infection (IMI) is often acute with local and systemic clinical manifestations that clear within 7 days. However, if not diagnosed early and treated, E. coli IMI could result in generalized systemic reaction and death. Persistent E. coli IMI is characterized by mild clinical manifestations followed by acute episodes of clinical mastitis during lactation. Factors responsible for pathogenesis of E. coli IMI and variation in clinical manifestations are not known. There are studies indicating that the outcome of E. coli IMI is mainly determined by cow factors. However, recent research demonstrated that virulence attributes of E. coli strains have significant impact on the outcome of E. coli IMI. The aims of this study were; (a) to compare gene expression profiles of PBMEC cocultured with strains of E. coli associated with acute or persistent IMI and; (b) to identify genes of E. coli induced during bacterial interaction with PBMEC. Utilizing cDNA we analyzed gene expression patterns of PBMEC cocultured with strains of E. coli using non-treated PBMEC as negative control. We evaluated also expression patterns of virulence associated genes of E. coli after co-culture with PBMEC using qRT-PCR. Our results showed that infection by both strains induced increased expression of pro-inflammatory cytokines, chemokines and innate immune response and apoptosis related genes. Our qRT-PCR results showed significant up-regulation of ler, eae, flic and iutA genes mainly in the strains of E. coli associated with persistent IMI. The pathogenesis and clinical severity of E. coli IMI may be determined by combined effects of host-pathogen factors.

The first reported case of human tick paralysis in Brazil: a new induction pattern by immature stages

Tick paralysis (TP) is a rare disease with rapid progression and potential fatal evolution. Immediately after the diagnosis, removal of all ticks from the body of the patient is mandatory. The present study reports for the first time a human case of the disease in Brazil. The patient had loss of muscle strength, decreased reflexes and marked palpebral ptosis. Six hours after removal of the last tick, the ptosis improved and on the following day, the patient had near total regression of the symptoms. This report emphasizes the possible presence of similar cases that should be promptly diagnosed and quickly treated. A new induction pattern for TP in humans associated with immature stages of ticks is also presented.

Organ donation and transplantation from medical students' perspective: introducing the experience from an academic league in Brazil.

INTRODUCTION: Medical education is an important factor to decrease the waiting list for transplantation. Reports in the medical literature reveal limited notification of potential organ donors by general physicians. Appropriate information is also needed to increase the availability of potential donors and minimize the waiting list. This article describes the acquired experience with an extracurricular program of education on organ and tissue transplantation in our institution, searching to meet medical information needs using a format of an academic league. METHODS: This qualitative study describes a proposed approach on the theme of "transplant and organ and tissue donation" with medical students from a Brazilian university, through creation of a program named "Transplantation League" in direct association with a transplantation center. RESULTS: The league's activities are based on three main activities teaching, research, and practical. Besides the organization of the I Course of Organ and Tissue Transplantation, the project received financial support from the Federal University of Goiás to develop the assignments. A member's stake in the league included scientific projects involving liver transplantation candidates, as well as notification, donation, transportation, and transplantation of these patients. CONCLUSION: The academic league has the purpose of academic information on organ and tissues transplantation. Its application in medical schools may be valuable to increase transplant numbers.

pGh9:ISS1 transpositional mutations in Streptococcus uberis UT888 causes reduced bacterial adherence to and internalization into bovine mammary epithelial cells.

Streptococcus uberis is an important mastitis pathogen that affects dairy cows worldwide. In spite of the economic impact caused by the high prevalence of S. uberis intramammary infections (IMI) in many well-managed dairy herds, pathogenic strategies and associated virulence factors of S. uberis are not well understood. It has been shown that S. uberis attaches to and internalizes into mammary epithelial cells and can survive inside cells for extended periods of time. We hypothesize that early attachment to and internalization into mammary epithelial cells is a critical step for the establishment of intramammary infection. The aim of this study is to identify and characterize chromosomally encoded virulence factors of S. uberis that allow early bacterial attachment to and internalization into mammary epithelial cells. A common approach used to identify virulence factors is by generating random insertion mutants that are defective in adherence to and internalization into mammary epithelial cells using pGh9:ISS1 mutagenesis system. A random insertion mutant library of S. uberis strain UT888 was created using a thermo-sensitive plasmid pGh9:ISS1 carrying ISS1 insertion sequence. Integration of the insertion sequence into the chromosome of these mutant clones was confirmed by PCR and Southern blot. Southern blot analysis of mutant clones also showed that insertional integration was random. Of 1000 random chromosomal insertion mutants of S. uberis strain UT888 screened, 32 had significantly reduced ability to adhere to and internalize into mammary epithelial cells. Chromosomal mapping of insertion sequence integration sites in some of these defective mutants showed integration into penicillin binding protein 2A (pbp2A), sensor histidine kinase, tetR family regulatory protein, phosphoribosylaminoimidazole carboxylase catalytic subunit (purE), lactose phosphotransferase, phosphoribosylamine glycine ligase (purD), and other genes involved in metabolic activities. These proteins may have a significant role in early bacterial colonization of the mammary gland during infection.

Presence of ISS1-like insertion sequence in wild type Streptococcus uberis strains isolated from cases of bovine mastitis.

Streptococcus uberis is a major cause of environmental mastitis worldwide. In spite of significant economic losses caused by S. uberis in many well-managed dairy herds, virulence factors and mechanisms associated with the pathogenesis of S. uberis mastitis are not well known. The ability of S. uberis to attach to and internalize into mammary epithelial cells and subsequent intracellular survival enables it to avoid host defense mechanisms. Research to determine virulence factors responsible for these pathogenic strategies involved creating a random chromosomal mutant library of S. uberis strain UT888 using the thermosensitive plasmid pGh9:ISS1 mutagenesis system. During Southern blot analysis of the mutant library, an endogenous element similar to ISS1 insertion sequence of Lactococcus lactis was found. ISS1 is a transposable bacterial insertion sequence isolated originally from L. lactis and are small phenotypically cryptic sequences of DNA with a simple genetic organization and capable of inserting at multiple sites in a target molecule. They are flanked by inverted repeats; generally encode their own transposition functions. A total of 29 of 34 wild type strains of S. uberis evaluated were positive for ISS1 by Southern blot. Insertion of ISS1 might have a significant phenotypic and genotypic role in the S. uberis genome because of its ability to transpose within the genome.

Vaccination of dairy cows with recombinant Streptococcus uberis adhesion molecule induces antibodies that reduce adherence to and internalization of S. uberis into bovine mammary epithelial cells.

Streptococcus uberis is an important environmental mastitis pathogen that causes subclinical and clinical mastitis in lactating and nonlactating cows and heifers throughout the world. Previous work from our laboratory suggests that S. uberis adhesion molecule (SUAM) is involved in S. uberis pathogenesis and may be an excellent target for vaccine development. The objective of this study was to evaluate the antibody response of cattle vaccinated with recombinant SUAM (rSUAM). Uninfected primiparous dairy cows (n=30) in late lactation were divided randomly into three groups of 10 cows each: control, 200 µg rSUAM, and 400 µg rSUAM. Cows in groups vaccinated with 200 µg and 400 µg rSUAM received an emulsion containing adjuvant, phosphate-buffered saline (PBS) and affinity purified rSUAM. Cows in the control group received an emulsion containing adjuvant and PBS. Cows were vaccinated subcutaneously in the neck region at drying off (D-0), 28 d after drying off (D+28) and within 7 d after calving. Serum was collected at D-0, D+28, at calving (C-0), calving vaccination (CV), and during early lactation (CV+14). Serum antibody responses were measured by an ELISA against rSUAM. Following the first vaccination a significant increase in anti-rSUAM antibodies was detected at D+28 in cows from groups vaccinated with 200 µg and 400 µg rSUAM when compared to the control group. This increase in anti-rSUAM antibodies continued following the second immunization at D+28; reaching the highest levels in the post-parturient sampling period (C0), after which antibodies appeared to plateau. S. uberis UT888 pretreated with several dilutions of heat-inactivated serum from cows vaccinated with rSUAM, affinity purified antibodies against rSUAM, and to a 17 amino acid long peptide from the N terminus of SUAM (pep-SUAM) were co-cultured with bovine mammary epithelial cells and adherence to and internalization of S. uberis into epithelial cells was measured. Compared to untreated controls, opsonization of two strains of S. uberis with sera from cows vaccinated with rSUAM, with affinity purified rSUAM antibodies, or with affinity purified pep-SUAM antibodies significantly reduced adherence to and internalization of this pathogen into bovine mammary epithelial cells. In conclusion, subcutaneous vaccination of dairy cows with rSUAM during physiological transitions of the mammary gland either from or to a state of active milk synthesis induced antibodies in serum and milk and these antibodies reduced adherence to and internalization of S. uberis into mammary epithelial cells under in vitro conditions. SUAM appears to be an excellent candidate for vaccine development.

Predicted antigenic regions of Streptococcus uberis adhesion molecule (SUAM) are involved in adherence to and internalization into mammary epithelial cells.

Streptococcus uberis is a significant cause of bovine mastitis throughout the world. Previous work from our laboratory demonstrated that S. uberis adhesion molecule (SUAM) is an important factor in adherence to and internalization of S. uberis into bovine mammary epithelial cells. Antibodies directed against SUAM significantly reduced bacterial adherence to and internalization into bovine mammary epithelial cells implying that SUAM is surface exposed. Objectives of this research were to: (1) predict surface exposed peptides, and (2) select peptide sequences for production of synthetic peptides with the final aim of evaluating their role in adherence and internalization and immunogenic potential. The Kyte/Doolittle hydropathicity prediction method; Chou/Fasman ß-turn prediction method; and output from Coils, Paircoil and MultiCoil scores for prediction of secondary and tertiary structures were used. Prediction algorithms resulted in identification of five overlapping regions of the SUAM sequence with the most hydrophilic valleys and the highest peaks for ß-turns. The five 15-mer SUAM epitopes selected by bioinformatic analysis were produced to evaluate the immunogenic value and pathogenic role of these putative domains. Peptides were bound to fluorescent latex beads, incubated with MAC-T bovine mammary epithelial cells, and internalization into MAC-T cells was evaluated using confocal laser and transmission electron microscopy. All peptides evaluated induced some degree of internalization of fluorescent beads into MAC-T cells; however, 2 peptides induced significantly more internalization of fluorescent beads than the other peptides evaluated. These peptides, designated III and IV, were located in the central region of SUAM, between two coiled-coil regions. Convalescent sera were tested against these biotinylated peptides for SUAM specific immune response using an indirect ELISA format. Among the 5 peptides evaluated, peptides I, II and V elicited significant serological response suggesting that the N-terminal region (peptide I), central region (peptide II) and C-terminal region (peptide V) are immunodominant epitopes of SUAM. Results will be useful to design immunotherapeutic tools based on immunodominant epitopes.

Opinion of hepatic transplant waiting list patients regarding split liver transplantation.

BACKGROUND: Split liver transplantation (SLT) increases organ supply for hepatic transplantation. Long-term patient survival and complication rates seem to be equivalent between orthotopic liver transplantation (OLT) and SLT. There are controversies among transplant physicians due to an ethical dilemma between benefiting individual needs or those of society. Barshes and Goss (Am J Transplant 5:2047, 2005) demonstrated that the majority of adult liver transplant candidates are favorable to SLT. The aim of our study was to evaluate the opinions of patients at a Brazilian university hospital regarding SLT. MATERIALS AND METHODS: A questionnaire with 14 questions was applied to 50 patients included in a hepatic transplant waiting list regarding SLT. RESULTS: The overall attitudes of 66% of the participants were classified as utilitarian, 31% were classified as self-preserving, and 3% were undecided. Ninety-one percent of patients would be willing to share even if their expected survival after SLT was shorter than that with OLT. For 77% of patients, children must have priority over adults. However, 83% were unaware of the donors for pediatric transplantations. CONCLUSIONS: SLT is a consistent solution for organ demand despite controversies among transplant physicians. The present study demonstrated that most patients were favorable to SLT. In conclusion, attitudes toward graft sharing are not barriers to SLT.

Assessment of cytokine values in serum by RT-PCR in HIV-1 infected individuals with and without highly active anti-retroviral therapy (HAART)

A cross-sectional study was performed on HIV-1 infected individuals with or without antiretroviral treatment (ARV) in the AIDS Day Hospital, Botucatu Medical School, UNESP. Between August 2004 and October 2005, 73 HIV-1 infected individuals were divided into three groups: infected individuals with or without AIDS who had never received ARV (G1 = 15); patients on HAART that had had plasma HIV-1 RNA viral load (VL) equal to or greater than 50 copies/mL (G2 = 27); and patients on HAART with undetectable VL for at least the past six months (G3 = 31). There was also an additional group that comprised blood donors without any sign of the disease and with negative HIV serum tests (G4 = 20), which was the control group. Serum cytokine levels (values in pg/mL) were measured by enzyme-linked immunosorbent assay (ELISA) and specific mRNA expression by reverse transcription polymerase chain reaction (RT-PCR). Both techniques were performed on the four groups for TNF-á, IL-2, INF-ã, IL-4 and IL-10. All patients were submitted to VL determination and CD4+ and CD8+T lymphocyte counts. The analysis of the results revealed a significant comparison among groups for both methods and an association between the latter (> 80% r² > 0.80). There was only one exception, in control individuals for IL-2 by ELISA. The cytokine profiles, in both methods, for the three patient groups, were mature Th-0. The behaviors of IL-2 and INF-ã required emphasis due to consequent expression of dominant Th profile. Both methods showed low IL-2 and high mean values of INF-ã in the three groups. Several authors have recently drawn attention to the substantial apoptosis of infected and non-infected CD4+T cells, mainly during primary infection, persisting only in those with INF-ã phenotype producer and not IL-2. HIV infected individuals submitted to HAART are expected to produce IL-2 in an attempt to present Th-1 profile, but in most cases this did not occur.(AU)

Discriminating between strains of Escherichia coli using pulsed-field gel electrophoresis and BOX-PCR.

In this study, we evaluated two biomolecular techniques for discriminating between strains of Escherichia coli isolated form a variety of sources. The DNA of 211 strains of E. coli collected from dairy farms, calves, feces, pigs, primates, humans, and food products was analyzed by pulsed-field gel electrophoresis (PFGE) and repetitive-element polymerase chain reaction using the BOXA1 primer (BOX-PCR). Objectives of the present study were to compare PFGE and BOX-PCR for discriminating among strains of E. coli and investigate their capability in clustering E. coli strains according to the origin of bacterial isolation. Our results showed that PFGE and BOX-PCR were both able to distinguish closely related strains of E. coli; however, PFGE was able to discriminate between isolates indistinguishable by BOX-PCR and interpretation of PFGE data was easier. BOX-PCR proved to have good discrimination power, was less expensive, and could be performed in a PCR thermocycler. Neither of the methods used were effective in clustering E. coli strains according to the source of the organism.

Intracellular killing of mastitis pathogens by penethamate hydriodide following internalization into mammary epithelial cells.

Penethamate hydriodide was highly effective in killing Streptococcus uberis, Streptococcus dysgalactiae subsp. dysgalactiae and Staphylococcus aureus that internalized mammary epithelial cells. At higher concentrations (32 microg/mL to 32 mg/mL), killing rates ranged from 85% to 100%. At lower concentrations (0.032 microg/mL to 3.2 microg/mL), killing rates ranged from 0 to 80%. Results of this proof-of-concept study demonstrated that: (1) penethamate hydriodide is capable of entering mammary epithelial cells and killing intracellular mastitis pathogens without affecting mammary epithelial cell viability, (2) the in vitro model used is capable of quantifying the fate of mastitis pathogens internalized into mammary epithelial cells, and (3) this in vitro model can be used to determine the effectiveness of antibiotics at killing bacteria within the cytoplasm of mammary epithelial cells.

Cytomegalovirus and Toxoplasma gondii seroprevalence in a Brazilian liver transplant waiting

Cytomegalovirus (CMV) disease is a major cause of morbidity and mortality in solid organ transplantation. Disseminated toxoplasmosis after liver transplantation is a rare but fatal event. Serologic screening of the donor and the recipient is essential to prophylactic management, early diagnosis and therapeutic strategies to minimize the consequences of these infections. The aim of the present study was to determine the seroprevalence of CMV and Toxoplasma gondii (TG) in a Brazilian liver transplant waiting list (LTWL). Serological data were collected from 44 candidates on the LTWL between May 2003 and November 2004. Serological investigation of antibodies IgM and IgG against CMV (anti-CMV) and TG (anti-T. gondii) was performed using fluorometry commercial kits. IgG anti-CMV was positive in 37 patients (94.9 percent) out of 39 available results. There were not IgM anti-CMV positive results. Out of 36 analyzed patients, 22 (61.1 percent) presented positive IgG anti-T. gondii and none had positive IgM anti-T. gondii. The high CMV seroprevalence among our LTWL reinforces the need for appropriate protocols to avoid related complications, like reactivation and superinfection by CMV. Environmental and drug prophylactic strategies against primary infection and reactivation, as well as early diagnosis and treatment of toxoplasmosis complications, are essential for the good outcome of transplant patients.

Serologic profile in hepatitis B virus and hepatitis C virus in a Brazilian liver transplant waiting list.

UNLABELLED: Chronic viral hepatitis is currently the most common indication for liver transplantation (OLT). Knowing the serological profile of patients on the liver transplant waiting list (LTWL) is essential to manage prophylactic and therapeutic strategies pre- and post-OLT. The aim of this study was to determine the hepatitis B virus (HBV) and hepatitis C virus (HCV) serological profile on the LTWL. METHODS: Serological data were collected from 44 candidates included on the LTWL from May 2003 to November 2004. HBV and HCV serological profiles were performed by microenzyme immunoassay. RESULTS: Twenty-eight patients (66.7%) lacked HBV serological markers. Anti-HBs was detected in 9.5% and was positive for HBsAg, anti-HBc, IgM anti-HBc, or HbeAg in 4.8% of patients, probably related to reactivation of chronic infection. In 7.1% of patients, the markers demonstrated serological cure of infection. In HCV patients, 41.5% were positive. There was HBV and HCV co-infection in 12.2% of patients. CONCLUSION: HBV infection in 21.4% of the patients corroborates the need to use more efficient protocols for prophylactic and therapeutic management pre- and post-OLT. The high prevalence of HCV infection reinforces the need to follow adequate protocols to avoid related complications and guarantee rational and universal use of more efficient drugs.

Adherent and invasive Escherichia coli are associated with persistent bovine mastitis.

Bovine mastitis caused by Escherichia coli has traditionally been viewed as a transient infection. However, E. coli can also cause clonal persistent intramammary infection (IMI) in dairy cows. In this study, we explored the possibility that E. coli strains associated with persistent IMI are better able to adhere to, invade, survive and replicate in cultured mammary epithelial cells (MAC-T) than transient strains, and examined their serotype, overall genotype, phylogenetic group, and the presence of known virulence genes. Both transient and persistent E. coli strains adhered to MAC-T cells, but persistent strains invaded MAC-T cells 2.6-63.5 times more than transient strains. Blocking the adhesin/invasin FimH with mannose diminished but did not eliminate adhesion and invasion of any strain. Cytoskeletal and protein kinase inhibitors cytochalasin D, colchicine, genistein and wortmannin dramatically reduced invasion of MAC-T cells by both strains. All of the persistent strains, but only one transient strain, were able to survive and replicate intracellularly in MAC-T cells over 48 h. Transient and persistent strains displayed heterogeneous serotypes and overall genotypes, but similar phylogeny (group A), and lacked virulence genes of invasive E. coli. We have found that E. coli strains associated with persistent IMI are better able to invade and replicate within cultured mammary epithelial cells than transient strains. The invasion process involves the host cytoskeleton and signaling cascades and is not FimH dependent. Our findings suggest that the invasion of mammary epithelial cells and intracellular survival play an important role in the pathogenesis of persistent E. coli mastitis.