Role of enhanced detoxication in a deltamethrin-resistant population of Triatoma infestans (Hemiptera, Reduviidae) from Argentina

Deltamethrin and other pyrethroids have been extensively used in Argentina since 1980, for the chemical control of Triatoma infestans Klug (Hemiptera: Reduviidae). Recently, resistance to deltamethrin was detected in field populations by the survival of bugs exposed by topical application to the diagnostic dose estimated on the CIPEIN susceptible strain. Results of the current study showed low resistant ratios (RRs) to deltamethrin for the resistant populations (RR ranged from 2.0 for San Luis colony to 7.9 for Salta colony). Biochemical studies were made on the most resistant colony (Salta) and the susceptible strain (CIPEIN), in order to establish the importance of degradative mechanisms as a cause of the detected resistance. Esterase activity was measured on 3 days old first instars through phenylthioacetate and a-naphtyl acetate activities. The results showed a significant difference in no cholinesterase esterase activity from susceptible (7.6 ± 0,7 µM S./i.min.) and Salta resistant colony (9.5 ± 0.8 µM S./i.min.). Cytochrome P450 mono-oxygenase (P450) activity was measured on individual insects through ethoxycoumarine deethylase (ECOD) activity using a fluorescence micro plate reader. The dependence of ECOD activity on age and body region of the nymphs, and pH and time of incubation were studied in order to optimize the measurement. As a result, comparative studies were performed on abdomens of 2 days old first instars at pH 7.2 and 4 h incubation time. ECOD activity of first nymphs was significantly lower in the susceptible colony (61.3 ± 9.08 pg ECOD/ insect) than in the resistant one (108.1± 5.7 pg ECOD/ insect). These results suggest that degradative esterases (no-cholinesterase) and mono-oxygenases cytochrome P450, play an important role in the resistance to deltamethrin in Salta colony from Argentina.

Antimicrobial resistance of Enterococcus sp. isolated from the intestinal tract of patients from a university hospital in Brazil

This study reports the results about antimicrobial resistance of Enterococcus spp. isolated from intestinal tract of patients from a university hospital in Brazil. The identification of strains at species level was performed by conventional biochemical tests, API 20 Strep (bioMérieux), and polymerase chain reaction assay. The specie distribution was E. faecium (34%), followed by E. faecalis (33%), E. gallinarum (23.7%), E. casseliflavus (5.2%), E. avium (1%), and E. hirae (1%). Intrinsic resistance to vancomycin characterized by presence of vanC genes was found in E. gallinarum and E. casseliflavus. The high prevalence of VanC phenotype enterococci is very important because these species have been reported as causing a wide variety of infections. Vancomycin-resistant E. faecium or E. faecalis were not found and no one isolate of these species was a beta-lactamase producer. Thirteen clinical isolates of enterococci (13.4%) showed multiresistance patterns, which were defined by resistance to three classes of antibiotics plus resistance to at least one aminoglycoside (gentamicin and/or streptomycin). The resistance to several antimicrobials shown by enterococcal strains obtained in this study is of concern because of the decrease in the therapeutic options for treatment of infections caused by enterococci.

Susceptibility of clinical isolates of multiresistant Pseudomonas aeruginosa to a hospital disinfectant and molecular typing

The aim of this study was to evaluate the susceptibility of 35 resistant Pseudomonas aeruginosa clinical isolates to a quaternary ammonium hospital disinfectant. The methodology was the AOAC Use-Dilution Test, with disinfectant at its use-concentration. In addition, the chromosomal DNA profile of the isolates were determined by macro-restriction pulsed field gel electrophoresis (PFGE) method aiming to verify the relatedness among them and the behavior of isolates from the same group regarding the susceptibility to the disinfectant. Seventy one percent of the isolates were multiresistant to antibiotics and 43% showed a reduced susceptibility to the disinfectant. The PFGE methodology detected 18 major clonal groups. We found isolates with reduced susceptibility to the disinfectant and we think that these are worrying data that should be further investigated including different organisms and chemical agents in order to demonstrate that microorganisms can be destroyed by biocide as necessary. We also found strains of the same clonal groups showing different susceptibility to the disinfectant. This is an interesting observation considering that only few works are available about this subject. PFGE profile seems not to be a reliable marker for resistance to disinfectants.

Toxicity of pyrethroids and repellency of diethyltoluamide in two deltamethrin-resistant colonies of Triatoma infestans Klug, 1834 (Hemiptera: Reduviidae)

The aim of the currrent investigation was to evaluate (a) the toxicity of three pyrethroids (deltamethrin, lambda-cyhalothrin, and tetramethrin); (b) the effect of these insecticides on the locomotor activity; and (c) the repellent effect of N,N-diethyl-m-toluamide (DEET) on two deltamethrin-resistant strains of Triatoma infestans from Argentina (El Chorro and La Toma), and one susceptible strain. The resistance ratios (RRs) obtained for the La Toma strain were: > 10,769, 50.7, and > 5.2 for deltamethrin, lambda-cyhalothrin, and tetramethrin respectively. The RRs for the El Chorro strain were: > 10,769, 85.8, and > 5.2 for deltamethrin, lambda-cyhalothrin, and tetramethrin respectively. The hyperactivity usually caused by the three pyrethroids was in both the deltamethrin-resistant strains compared to the susceptible reference strain. No differences were observed in the repellent effect of DEET between the three groups. These results indicate that the deltamethrin-resistant insects have a cross resistance to lambda-cyhalothrin and tetramethrin, and are also resistant to the first symptom of pyrethroid poisoning (hyperactivity). However, the sensorial process related to DEET repellency does not appear to be altered.

Serogroups, K1 antigen, and antimicrobial resistance patterns of Aeromonas spp. strains isolated from different sources in Mexico

A total of 221 strains of Aeromonas species isolated in Mexico from clinical (161), environmental (40), and food (20) samples were identified using the automated system bioMérieux-Vitek®. Antisera for serogroups O1 to 044 were tested using the Shimada and Sakazaki scheme. The K1 antigen was examined using as antiserum the O7:K1C of Escherichia coli. Besides, we studied the antimicrobial patterns according to Vitek AutoMicrobic system. Among the 161 clinical strains 60% were identified as A. hydrophila, 20.4% as A. caviae, and 19.25% as A. veronii biovar sobria. Only A. hydrophila and A. veronii biovar sobria were found in food (55 and 90% respectively) and environmental sources (45 and 10% respectively). Using "O" antisera, only 42.5% (94/221) of the strains were serologically identified, 55% (121/221) were non-typable, and 2.5% (6/221) were rough strains. Twenty-two different serogroups were found, O14, O16, O19, O22, and O34 represented 60% of the serotyped strains. More than 50% of Aeromonas strain examined (112/221) expressed K1 encapsulating antigen; this characteristic was predominant among Aeromonas strains of clinical origin. Resistance to ampicillin/sulbactam and cephazolin was detected in 100 and 67% of Aeromonas strain tested for their susceptibility to antibiotics. In conclusion, antibiotic-resistant Aeromonas species that possess the K1 encapsulating antigen and represent serogroups associated with clinical syndrome in man are not uncommon among Aeromonas strains isolated from clinical, food and environmental sources in Mexico.

In vitro susceptibility to antifungal agents of environmental Cryptococcus spp. isolated in the city of Ribeirão Preto, São Paulo, Brazil

Infections by Cryptococcus strains other than C. neoformans have been detected in immunocompromised patients. Of these strains, three are considered human pathogens: C. albidus, C. laurenttii, and C. uniguttulatus. This study deals with the in vitro susceptibility of Cryptococcus to drugs such as amphotericin B, itraconazole, fluconazole, and 5-fluorocytosine. Environmental Cryptococcus isolates (50) distributed as follows: C. neoformans var. neoformans (16), C. albidus (17), C. laurentii (14), and C. uniguttulatus (3) were evaluated by the micro and macrodilution techniques, according to EUCAST and NCCLS recommendations, respectively. Considering both methodologies the respective minimal inhibitory concentrations (MIC) were 0.125 and 2 µg/ml for amphotericin B, 0.06 and 8 µg/ml for itraconazole, and 0.5 and more than 64 µg/ml for fluconazole and 5-fluorocytosine. Agreement percentages for the two methodologies were 100% for amphotericin B and fluconazole for all the strains tested. For itraconazole, the agreement percentage was 81.3% in the C. neoformans strain and 100% for all the others. All species had a agreement percentage of 94.1 to 100% when susceptibility to 5-fluorocytosine was tested. It is concluded that environmental isolates of C. neoformans var. neoformans, C. albidus, C. laurentii, and C. uniguttulatus may show high MICs against certain drugs, suggesting in vitro primary resistance to the antifungals tested.

Primary resistance of human immunodeficiency virus type 1 in a reference center in Recife, Pernambuco, Brazil

To assess the prevalence of primary resistance of human immunodeficiency virus type 1 (HIV-1) to antiretrovirals, 84 patients chronically infected with HIV without prior antiretroviral treatment from Northeast Brazil were studied. Genotyping was performed using the ViroSeqTM Genotyping System. Thimidine analog mutations occurred in 3 (3.6%) patients. Accessory mutations related to NRTI occurred in 6 (7.1%) and related to PI in 67 (79.8%). Subtypes B (72.6%), F (22.6%), B/F 3 (3.6%), and C (1.2%) were detected. A low prevalence of major mutations related to NRTI in patients chronically infected by HIV was observed.

Differential lectin labelling of circulating hemocytes from Biomphalaria glabrata and Biomphalaria tenagophila resistant or susceptible to Schistosoma mansoni infection

Lectins/carbohydrate binding can be involved in the Schistosoma mansoni recognition and activation of the Biomphalaria hemocytes. Therefore, expression of lectin ligands on Biomphalaria hemocytes would be associated with snail resistance against S. mansoni infection. To test this hypothesis, circulating hemocytes were isolated from B. glabrata BH (snail strain highy susceptible to S. mansoni), B. tenagophila Cabo Frio (moderate susceptibility), and B. tenagophila Taim (completely resistant strains), labelled with FITC conjugated lectins (ConA, PNA, SBA, and WGA) and analyzed under fluorescence microscopy. The results demonstrated that although lectin-labelled hemocytes were detected in hemolymph of all snail species tested, circulating hemocytes from both strains of B. tenagophila showed a larger number of lectin-labelled cells than B. glabrata. Moreover, most of circulating hemocytes of B. tenagophila were intensively labelled by lectins PNA-FITC and WGA-FITC, while in B. glabrata small hemocytes were labeled mainly by ConA. Upon S. mansoni infection, lectin-labelled hemocytes almost disappeared from the hemolymph of Taim and accumulated in B. glabrata BH. The role of lectins/carbohydrate binding in resistance of B. tengophila infection to S. mansoni is still not fully understood, but the data suggest that there may be a correlation to its presence with susceptibility or resistance to the parasite.

Antiretroviral resistance in individuals presenting therapeutic failure and subtypes of the human immunodeficiency virus type 1 in the Northeast Region of Brazil

This study aimed to analyze human immunodeficiency virus (HIV) mutation profiles related to antiretroviral resistance following therapeutic failure, and the distribution of hiv subtypes in the Northeast Region of Brazil. A total of 576 blood samples from AIDS patients presenting therapeutic failure between 2002 and 2004 were analyzed. The genotyping kit viroSeq® was used to perform viral amplification in order to identify mutations related to hiv pol gene resistance. An index of 91.1% of the patients presented mutations for nucleoside reverse transcriptase inhibitors (nrti), 58.7% for non-nucleoside reverse transcriptase inhibitors (nnrti), and 94.8% for protease inhibitors (pi). The most prevalent mutations were 184V and 215E for nrti, 103N and 190A for nnrti. Most mutations associated with PIs were secondary, but significant frequencies were observed in codons 90 (25.2%), 82 (21.1%), and 30 (16.2%). The resistance index to one class of antiretrovirals was 14%, to two classes of antiretrovirals 61%, and to three classes 18.9%. Subtype B was the most prevalent (82.4%) followed by subtype F (11.8%). The prevalence of mutations related to nrti and nnrti was the same in the two subtypes, but codon analysis related to PI showed a higher frequency of mutations in codon 63 in subtype B and in codon 36 in subtype F. The present study showed that there was a high frequency of primary mutations, which offered resistance to nrti and nnrti. Monitoring patients with treatment failure is an important tool for aiding physicians in rescue therapy.

Antimicrobial resistance profiles of enterococci isolated from poultry meat and pasteurized milk in Rio de Janeiro, Brazil

The enterococci are important nosocomial pathogens with a remarkable capacity of expressing resistance to several antimicrobial agents. Their ubiquitous nature and resistance to adverse environmental conditions take account for their ability to colonize different habitats and for their potential for easy spreading through the food chain. In the present study we evaluated the distribution of species and antimicrobial susceptibility among enterococcal isolates recovered from food obtained in retail stores in Rio de Janeiro, Brazil. The following species were identified among 167 isolates obtained from poultry meat and 127 from pasteurized milk: Enterococcus faecalis (62.6%), E. casseliflavus (17.3%), E. durans (6.5%), E. gallinarum (3.0%), E. gilvus (2.4%), E. faecium (2.0%), E. hirae (1.4%), and E. sulfureus (1.0%). The overall percentages of antimicrobial resistant isolates were: 31.2 % to tetracycline, 23.8% to erythromycin, 11.3% to streptomycin, 4.3% to chloramphenicol, 3.9% to gentamicin, 1.4% to norfloxacin, 1.1% to imipenem, 0.7% to ciprofloxacin, nitrofurantoin, and penicillin, and 0.4% to ampicillin. Intermediate resistance was detected in frequencies varying from 0.5% for linezolid to 58.2% for erythromycin. None of the isolates showed resistance to glycopeptides. High-level resistance to aminoglycosides was observed in 13.1% of the isolates. Multiresistance was observed in E. faecalis, E. casseliflavus, E. faecium, E. gallinarum, E. durans and E. gilvus.

Antimicrobial resistance among Brazilian Corynebacterium diphtheriae strains

The increasing problems with multidrug resistance in relation to Corynebacterium, including C. diphtheriae, are examples of challenges confronting many countries. For this reason, Brazilian C. diphtheriae strains were evaluated by the E-Test for their susceptibility to nine antibacterial drugs used in therapy. Resistance (MIC < 0.002; 0.38 µg/ml) to penicillin G was found in 14.8% of the strains tested. Although erythromycin (MIC90 0.75 µg/ml) and azithromycin (MIC90 0.064 µg/ml) were active against C. diphtheriae in this study, 4.2% of the strains showed decreased susceptibility (MIC 1.0 µg/ml) to erythromycin. Multiple resistance profiles were determined by the disk diffusion method using 31 antibiotics. Most C. diphtheriae strains (95.74%) showed resistance to mupirocin, aztreonam, ceftazidime, and/or oxacillin, ampicillin, penicillin, tetracycline, clindamycin, lincomycin, and erythromycin. This study presents the antimicrobial susceptibility profiles of Brazilian C. diphtheriae isolates. The data are of value to practitioners, and suggest that some concern exists regarding the use of penicillin.

Added Value of Deep Sequencing Relative to Population Sequencing in Heavily Pre-Treated HIV-1-Infected Subjects.

OBJECTIVE: To explore the potential of deep HIV-1 sequencing for adding clinically relevant information relative to viral population sequencing in heavily pre-treated HIV-1-infected subjects. METHODS: In a proof-of-concept study, deep sequencing was compared to population sequencing in HIV-1-infected individuals with previous triple-class virological failure who also developed virologic failure to deep salvage therapy including, at least, darunavir, tipranavir, etravirine or raltegravir. Viral susceptibility was inferred before salvage therapy initiation and at virological failure using deep and population sequencing genotypes interpreted with the HIVdb, Rega and ANRS algorithms. The threshold level for mutant detection with deep sequencing was 1%. RESULTS: 7 subjects with previous exposure to a median of 15 antiretrovirals during a median of 13 years were included. Deep salvage therapy included darunavir, tipranavir, etravirine or raltegravir in 4, 2, 2 and 5 subjects, respectively. Self-reported treatment adherence was adequate in 4 and partial in 2; one individual underwent treatment interruption during follow-up. Deep sequencing detected all mutations found by population sequencing and identified additional resistance mutations in all but one individual, predominantly after virological failure to deep salvage therapy. Additional genotypic information led to consistent decreases in predicted susceptibility to etravirine, efavirenz, nucleoside reverse transcriptase inhibitors and indinavir in 2, 1, 2 and 1 subject, respectively. Deep sequencing data did not consistently modify the susceptibility predictions achieved with population sequencing for darunavir, tipranavir or raltegravir. CONCLUSIONS: In this subset of heavily pre-treated individuals, deep sequencing improved the assessment of genotypic resistance to etravirine, but did not consistently provide additional information on darunavir, tipranavir or raltegravir susceptibility. These data may inform the design of future studies addressing the clinical value of minority drug-resistant variants in treatment-experienced subjects.

Mixed-function oxidases and esterases associated with cross-r esistance between DDT and lambda-cyhalothrin in Anopheles darlingi Root 1926 populations from Colombia

In order to establish the insecticide susceptibility status for Anopheles darlingi in Colombia, and as part of the National Network on Insecticide Resistance Surveillance, five populations of insects from three Colombian states were evaluated. Standardised WHO and CDC bottle bioassays, in addition to microplate biochemical assays, were conducted. Populations with mortality rates below 80% in the bioassays were considered resistant. All field populations were susceptible to deltamethrin, permethrin, malathion and fenitrothion. Resistance to lambda-cyhalothrin and DDT was detected in the Amé-Beté population using both bioassay methods with mortality rates of 65-75%. Enzyme levels related to insecticide resistance, including mixed function oxidases (MFO), non-specific esterases (NSE), glutathione S-transferases and modified acetylcholinesterase were evaluated in all populations and compared with a susceptible natural strain. Only mosquitoes from Amé-Beté presented significantly increased levels of both MFO and NSE, consistent with the low mortalities found in this population. The continued use of lambda-cyhalothrin for An. darlingi control in this locality has resulted in a natural resistance to this insecticide. In addition, DDT resistance is still present in this population, although this insecticide has not been used in Colombia since 1992. Increased metabolism through MFO and NSE may be involved in cross-resistance between lambda-cyhalothrin and DDT, although kdr-type nerve insensitivity cannot be discarded as a possible hypothesis. Additional research, including development of a kdr specific assay for An. darlingi should be conducted in future studies. Our data demonstrates the urgent need to develop local insecticide resistance management and surveillance programs throughout Colombia.

Advances in understanding immunity to Toxoplasma gondii

Toxoplasma gondii is an important cause of clinical disease in fetuses, infants and immunocompromised patients. Since the discovery of T. gondii 100 years ago, this pathogen and the host's immune response to toxoplasmosis have been studied intensely. This has led to the development of a working model of immunity to T. gondii, and has also resulted in fundamental new insights into the role of various cytokines in resistance to infection. By examining this organism, researchers have identified many of the requirements for resistance to intracellular pathogens and characterized numerous regulatory factors, including interleukin-10 (IL-10) and IL-27, which control inflammatory processes. In the next 100 years of T. gondii immunobiology, researchers will have the opportunity to answer some of the long-standing questions in the field using new techniques and reagents. These future studies will be vital in building a more comprehensive model of immunity to this pathogen and in advancing our understanding of immunoregulation, particularly in humans. Ultimately, the challenge will be to use this information to develop new vaccines and therapies to manage disease in affected patients.

Toxoplasma gondii and the Immunity-Related GTPase (IRG) resistance system in mice: a review

The Immunity Related GTPases (IRG proteins) constitute a large family of interferon-inducible proteins that mediate early resistance to Toxoplasma gondii infection in mice. At least six members of this family are required for resistance of mice to virulent T. gondii strains. Recent results have shown that the complexity of the resistance arises from complex regulatory interactions between different family members. The mode of action against T. gondii depends on the ability of IRG proteins to accumulate on the parasitophorous vacuole of invading tachyzoites and to induce local damage to the vacuole resulting in disruption of the vacuolar membrane. Virulent strains of T. gondiiovercome the IRG resistance system, probably by interfering with the loading of IRG proteins onto the parasitophorous vacuole membrane. It may be assumed that T. gondii strains highly virulent for mice will be disadvantaged in the wild due to the rapid extinction of the infected host, while it is self-evident that susceptibility to virulent strains is disadvantageous to the mouse host. We consider the possibility that this double disadvantage is compensated in wild populations by segregating alleles with different resistance and susceptibility properties in the IRG system.