Multiple insecticide resistance in Aedes aegypti (Diptera: Culicidae) from Boyeros municipality, Cuba and associated mechanisms.

In this study, insecticide resistance and the mechanisms responsible were characterized in Ae. aegypti of Boyeros municipality from Havana, Cuba. Boyeros represents a high epidemiological risk because it is located near the Havana International Airport, it is highly urbanized, and it has a large influx of people from endemic countries so that it qualifies as a sentinel site for surveillance. The larvae collected from five areas of this municipality showed resistance to temephos associated with metabolic enzymes. The adult mosquitoes displayed a deltamethrin resistance and less distinctly to other pyrethroids associated with a high frequency of sodium channel gene mutations (F1534C and V1016I), detected for the first time in a field population from Cuba. The presence in the field populations of two insecticide resistance mechanisms represents a limiting factor in the success of the control operations of this vector, so other strategies should be considered to preserve the effectiveness of the insecticides available in public health for vector control in Cuba.

Insecticide resistance in two Aedes aegypti (Diptera: Culicidae) strains from Costa Rica.

Dengue (family Flaviridae, genus Flavivirus, DENV) and dengue hemorrhagic fever (DHF) are presently important public health problems in Costa Rica. The primary strategy for disease control is based on reducing population densities of the main mosquito vector Aedes aegypti (L.) (Diptera: Culicidae). This is heavily dependent on use of chemical insecticides, thus the development of resistance is a frequent threat to control program effectiveness. The objective of this study was to determine the levels of insecticide resistance and the metabolic resistance mechanisms involved in two Ae. aegypti strains collected from two provinces (Puntarenas and Limon) in Costa Rica. Bioassays with larvae were performed according to World Health Organization guidelines and resistance in adults was measured through standard bottle assays. The activities of beta-esterases, cytochrome P450 monooxygenases, and glutathione S-transferases (GST), were assayed through synergists and biochemical tests, wherein the threshold criteria for each enzyme was established using the susceptible Rockefeller strain. The results showed higher resistance levels to the organophosphate (OP) temephos and the pyrethroid deltamethrin in larvae. The efficacy of commercial formulations of temephos in controlling Ae. aegypti populations was 100% mortality up to 11 and 12 d posttreatment with daily water replacements in test containers. Temephos and deltamethrin resistance in larvae were associated with high esterase activity, but not to cytochrome P450 monooxygenase or GST activities. Adult mosquitoes were resistant to deltamethrin, and susceptible to bendiocarb, chlorpyrifos, and cypermethrin. Because temephos and deltamethrin resistance are emerging at the studied sites, alternative insecticides should be considered. The insecticides chlorpyrifos and cypermethrin could be good candidates to use as alternatives for Ae. aegypti control.

Short-lived corpora lutea syndrome in anoestrous ewes following 17ß-oestradiol or MAP treatments applied before an allogenic sexual stimulation with rams and oestrous ewes.

When induced to ovulate during anoestrus, ewes, does and cows frequently develop a short-lived corpora lutea (SLCL) syndrome associated to lack of previous progesterone. Exogenous progesterone precludes SLCL by blocking oxytocin endometrial receptors, thus inducing normal life-span CL (NLCL). Paradoxically, circa 50% of unprimed ewes do not develop SLCL. We report results from 3 trials assessing follicular, oestrous, ovulatory, and luteal end-points after 17ß-oestradiol or MAP treatments. Oestradiol benzoate (50µg) induced follicular turnover, provoked ovulation in 40% (24/60) of ewes treated (93% of which developed SLCL), but did not affect the incidence of SLCL (26/53) after an allogenic sexual stimulation (ASS) by rams and oestrous ewes. By the onset of the ASS, most NLCL ewes (26/27) had already experienced turnover of their largest follicle, had smaller largest and second largest follicles, and ovulated their largest follicle more frequently than SLCL ewes did. Most SLCL ewes (19/25) did not ovulate their largest follicles, ovulating instead smaller follicles of identical size to those of NLCL ewes. Priming (40mg of MAP for 12 days) was partially effective at preventing SLCL even when terminated 14 days in advance of an ASS, but failed at completely preventing SLCL when terminated 6 or more days in advance. The coupling of a timed acquisition of full steroidogenic capability before ovulation with a system of endometrial oestradiol-progesterone-oxytocin receptors linked in an unstable equilibrium controlling the amplification of the luteolytic feed-forward loop of oxytocin and prostaglandin F(2)α explains occurrence and relative incidences of both NLCL and SLCL, and links proximate and ultimate causes of the SLCL syndrome.

Temephos resistance and esterase activity in the mosquito Aedes aegypti in Havana, Cuba increased dramatically between 2006 and 2008.

Aedes aegypti (L.) (Diptera: Culicidae) control programmes in Cuba rely on the application of the organophosphate temephos for larval control. Hence, the monitoring of resistance to this insecticide is an essential component of such programmes. Here, 15 field populations from different municipalities of Havana City were assayed for resistance to temephos. High levels of resistance were detected in all strains and resistance ratios were highly correlated with esterase activity (P = 0.00001). Populations from three municipalities were tested in both 2006 and 2008; resistance and esterase activities both significantly increased during this 2-year period. Synergist studies demonstrated that neither glutathione transferases nor monooxygenases were associated with the increase in resistance to temephos in this period. The duration of the efficacy of commercial formulations of temephos in controlling Ae. aegypti populations in Havana City was reduced by the high level of temephos resistance observed; hence these data are of clear operational significance for the dengue control programme in Cuba. New integrated strategies to avoid further increases in temephos resistance in Cuba are necessary.

Relación de los factores de riesgo y comorbilidad en el programa de prevención de caídas en usuarios hospitalizados en un hospital psiquiátrico del sector salud / Ratio risk factors and comorbidity in the program prevention of falls in users hospitalized in a hospital mental health sector

Introducción: Los pacientes hospitalizados en una unidad de salud mental tienen el riesgo potencial de enfrentar factores de riesgo. Con la intensión de prevenir caídas en los usuarios hospitalizados Ruelas y Sarabia (10, 11), han elaborado, un modelo de valoración de los factores de riesgo como son: falta iluminación adecuada, desorden del mobiliario, falta de limpieza del área física, desconocimiento de las medidas de seguridad dentro de las instalaciones de la unidad, así como factores inherentes al paciente y su condición como son: sexo, edad, estilo de vida, movilidad, factores relacionados con el estado físico, sensoriales, estado neurológico y emocional alterados, tratamiento farmacológico y los relacionados con los dispositivos para deambular. Material y Método: Se realizó una investigación de tipo transversal y descriptiva, de Mayo del 2008 a Mayo del 2009, la muestra la conformaron 285 pacientes hospitalizados a los cuales se les aplicó una cédula de evaluación de riesgo de caídas al ingreso y cada 7 días hasta su egreso. Resultados: Durante el tiempo de hospitalización 5% de los pacientes presentaron caída, entre las características de estos tenemos el ser mujer 76%, recibir tratamiento con benzodiacepinas (clonazepan) 100%, con diagnóstico de ingreso episodios depresivos graves con pensamiento o intento suicida 53%, con conducta disruptiva, por episodio maniaco o trastorno psicótico 38%, con reporte de resonancia magnética de pérdida de volumen de parénquima cerebral 61% y factores extrínsecos 28%. Conclusión: Se identificó que los factores de riesgo predominantes en nuestros pacientes son la combinación de psicofármacos, y la enfermedad mental, la falta de identificación del riesgo en pacientes, familiares, cuidadores y equipo de salud. En menor porcentaje influyen los factores estructurales.

Introduction: Hospitalized patients in a Mind Health Unit face factors of risk.Some authors (10, 11) have made a model to value the factors for preventing falling such as: inadequate lighting, lack of placement of furniture, lack of cleaning the physical area, lack of security measures and patient characteristics like: sex, age, life style, mobility, fitness-related factors, sensory factors, being emotional and neurological altered pharmacology treatment and related devices for wandering. Material and Methods: This is a descriptive, and transversal research realized from May 2008 to May 2009, a card falling schedule was applied to 285 patients since their income to the mind health unit and every 7 days until their discharge. Results: During the hospitalization time 5% of patients were falling; 76% were women, 100% with benzodiazepines (clonazepan) treatment, 53% with depressing episodes and suicide attempts, 38% with disruptive conduct for manic episode or psychotic disorder, 61% with loss brain parenchyma volume and 28% for extrinsic factors. The most frequently risk factors founded in our patients are the combination of drugs and mental illness, unknown ledge of risk by patients, their family, their careers, and health team. Structural factors risks have the lower percentage.

LH response of seasonally anovular Corriedale ewes acutely exposed to rams and estrous ewes.

Serial blood sampling before and after exposing four anovular Corriedale ewes to a group of rams and estrous ewes during the non-breeding season revealed a pattern of LH secretion similar to that previously observed in Merinos. Mean LH values doubled (P<0.001) from 0.24+/-0.06 microgL(-1) (mean+/-s.e.m.) before to 0.55+/-0.05 microgL(-1) after 2h of visual, auditory, and odor exposure to rams and estrous ewes in an indoor facility. A non-significant (P<0.17) increase of LH pulses per hour was also observed (0.7+/-0.3 pulses per hour before compared with 1.3+/-0.3 during stimulation). All four ewes had recently formed corpora lutea by five days after stimulation. Results are consistent with the pattern of sudden increase and sustained release of LH observed in other sheep breeds, particularly the Merino.

Immunobiological role of llama heavy-chain antibodies against a bacterial beta-lactamase.

In 1993, a fraction of antibodies (Abs) devoid of L chain was found naturally occurring in the Camelidae. They were found to lack L chains, as well as the first constant heavy-chain domain (CH(1)) and therefore they were named "heavy-chain Abs" (HCAbs). Subsequent studies focused on the functional, structural and biochemical properties of recombinant variable fragments (rVHHs) of HCAbs. It was stated that rVHHs have an augmented capacity to interact with "partially hidden" epitopes, like enzymes active sites, and have an increased stability to thermal and chemical aggression. It has been suggested that these unconventional Abs could represent an evolutionary advantage, being more efficient than conventional Abs to inhibit microbial enzymes, and thus exerting a more protective immune response against pathogens. The present work focuses on the immunobiological role of HCAbs, in their capacity to inhibit microbial enzymes. Two animal models were selected, comprising a model for common vertebrates without HCAbs (rabbits), and a model for vertebrates with both conventional and unconventional Abs (Lama glama). A recombinant bacterial beta-lactamase (CTX-M-2) was selected as the microbial enzymatic antigen. After conventional immunization schedules, neither serum titers nor serum inhibitory capacity showed significant differences when rabbits and llamas were compared. These results indicate that the a priori assumption that the adaptive immune system of camelids could be better "prepared" to respond to bacterial enzymes because of the presence of HCAbs, is not always accurate. Furthermore, when the different llama antibody isotypes and subclasses were purified, it was demonstrated that the inhibitory capacity of total serum was due exclusively to IgG(1). HCAbs not only failed to inhibit CTX-M-2, but instead they activated its enzymatic activity. Altogether, these results indicate that the hypotheses extrapolated from the rVHHs properties need to be revised; the real role of HCAbs in vivo remains unknown, as well as their evolutionary cause.

Application of the pupal/demographic-survey methodology in an area of Havana, Cuba, with low densities of Aedes aegypti (L.).

Between the April and December of 2004, an Aedes aegypti (L.) pupal/demographic survey was conducted in the Playa municipality of Havana. The aims were to identify the key types of container in the production of the adult mosquitoes (and so provide a basis for targeted control) and assess transmission risk in terms of the number of pupae/person. Pupal abundance, as measured in the survey, was compared with traditional Stegomyia indices. The immature stages of Ae. aegypti were only found in 70 of the 15,153 containers that were investigated and the pupae of this species were only seen in 52 of the containers. Ground-level water-storage tanks yielded 74.1% of all the pupae, with a further 19.0% found in miscellaneous small containers. The utility of the pupal/demographic-survey methodology in the evaluation of dengue risk in the study area, and other areas of Cuba with low densities of Ae. aegypti, is discussed.

A novel mutation (G114V) in the prion protein gene in a family with inherited prion disease.

Inherited prion diseases are characterized by mutations in the PRNP gene encoding the prion protein (PrP). We report a novel missense mutation in the PRNP gene (resulting in a G114V mutation in PrP) in members of a Uruguayan family with clinical and histopathologic features of prion disease. Affected individuals were characterized by an early age at onset, initial neuropsychiatric symptoms, late dementia with prominent pyramidal and extrapyramidal symptoms, and long disease duration.

(GCG)11 founder mutation in the PABPN1 gene of OPMD Uruguayan families.

The dominant oculo-pharyngeal muscular dystrophy mutation consists of an expanded (GCN)(12-17) in the coding region of the PolyA Binding Protein Nuclear 1 gene. A founder effect has been demonstrated in Canadian and Bukhara Jewish populations with relatively high prevalence of this disease. Since the oculo-pharyngeal muscular dystrophy prevalence was remarkably high in Southern Uruguay, a founder effect was hypothesized. To identify the ancestral haplotype we determined the (GCN) repeat number and the variants of four intragenic SNPs in Uruguayan OPMD families and a control sample. All families carrying the mutation (GCG)(11)(GCA)(3)(GCG) shared a common ancestral haplotype and the age of the mutation was estimated in 37-53 generations by a composite likelihood method. One family carrying the (GCG)(9)(GCA)(3)(GCG) allele had a different haplotype. The genealogical and molecular data suggested that the common ancestors were Canary Islands' settlers that arrived in Uruguay in the XIX century.

Extended-spectrum beta-lactamases in enterobacteriaceae in Buenos Aires, Argentina, public hospitals.

Resistance to extended-spectrum cephalosporins is often associated with plasmid encoded extended spectrum beta-lactamases (ESBL). In order to evaluate the prevalence and diversity of ESBLs in enterobacteria in our city, a 1-month-period survey was carried out from April to May 2000. Extended-spectrum-cephalosporin-resistant strains, isolated from inpatient clinical specimens other than stools, were collected among 17 participating hospitals. From a total of 427 enterobacterial strains that were collected during this period, 39 were extended-spectrum cephalosporin resistant. The National Committee for Clinical Laboratory Standards' Screening and Confirmatory Tests for ESBL production were performed using cefotaxime and ceftazidime; cefepime and cefepime-clavulanic acid-containing disks were included. beta-Lactamases were characterized by isoelectric focusing and PCR amplification using specific primers. Three different ESBLs were detected: SHV-related (4 isolates), PER-2-type (9 isolates), and CTX-M-2-related (26 isolates). Sequencing of the corresponding genes confirmed CTX-M-2 in 19 of 21 and CTX-M-31 (an allelic variant) in the remaining 2 of 21. CTX-M-2 (or its variant) was detected in all Escherichia coli, Enterobacter aerogenes, Serratia marcescens, Proteus mirabilis, and Providencia stuartii strains, while PER-2 was detected in Enterobacter cloacae, E. aerogenes, and Klebsiella pneumoniae; SHV-related ESBL were found only in K. pneumoniae. These results clearly show that CTX-M-2 is the most prevalent ESBL produced by enterobacterial species isolated from public hospitals in Buenos Aires.

Detection of insecticide resistance in Aedes aegypti (Diptera: Culicidae) from Cuba and Venezuela.

Four strains of Aedes aegypti (L.), one from Cuba and three from Venezuela, were bioassayed for susceptibility to eight insecticides, including the organophosphates, temephos, malathion, fenthion, pirimiphos methyl, and chlorpyrifos, and the pyrethroids, deltamethrin, lambda cyhalothrin and cypermethrin, S, S, S,-tributyl phosphorotrithioate and piperonyl butoxide were used as synergists to assess the involvement of esterases and monooxygenases in organophosphate resistance. Venezuelan strains had low levels of resistance to fenthion and malathion, and moderate to high resistance to temephos, pyrimphos methyl, and chlorpiriphos. All strains were susceptible to the pyrethroids, except the Cuban strain, which had moderate levels of resistance to cypermethrin. Organophosphate resistance in Ae. aegypti is a serious threat to control operations. Integrated strategies for Ae. aegypti control to prevent or delay pyrethroid resistance in Venezuela and Cuba are discussed.

[High esterases as mechanism of resistance to organophosphate insecticides in Aedes aegypti strains]. / Esterasas elevadas como mecanismo de resistencia a insecticidas organofosforados en cepas de Aedes aegypti.

Five strains of Aedes aegypti L. one from Cuba and 4 from Venezuela were evaluated to determine their resistance to organophosphate insecticides (temephos, chlorpiriphos and pirimiphos methyl). In the Venezuelan strains only APURE showed resistance to temephos. In TACHIRA and MIRANDA moderate resistance values were noted (FR50 5 to 10x) for chlorpiriphos and high levels of resistance (FR > 10x) to this insecticide were found in ARAGUA. All the Venezuelan strains showed high levels of resistance to pirimiphos methyl. The Cuban strain from Santiago de Cuba revealed moderate resistance to temephos and pirimiphos methyl, but high resistance to chlorpiriphos. The results of the biochemical tests proved the presence of esterase and glutathione-s-transferase at high frequency in almost every strain. By the polyacrilamide gel electrophoresis, a strongly stained band was observed in all the strains with a Rf value of 0.779; it was named esterase A4 and was not seen in the susceptible reference strain. The meaning of this esterase in the resistance to organophosphate insecticides is yet to be determined. Resistance to these insecticides in Aedes aegypti is a serious problem for the control of this species therefore integrated management strategies were proposed to prevent or delay the appearance of this species in Cuba and Venezuela.

[Adaptation of microtitration plate methods for quantification of the activity of esterases and glutathione-s-transferase in Aedes aegypti]. / Adaptación de los métodos en placas de microtitulación para la cuantificación de la actividad de esterasas y glutation-s-transferasa en Aedes aegypti.

The detection techniques for the activity of non-specific esterases and glutathione-s-transferase in Culex quinquefasciatus were modified to detect such enzymes in Aedes aegypti(L). The optimal concentration values of substrate (saturating concentration) and the optimum reading time for reaction were determined for each enzyme by using 4 Aedes aegypti strains: one from Cuba 2 from Venezuela and one susceptible reference strain. The frequency of non-specific esterases turned out to be 0.76 in MIRANDA 0.42 in ARAGUA and 1 in SANTIAGO DE CUBA in which the highest frequency value of this mechanisms was reached. The frequency of glutathione-s-transferase mechanism was 0.45 in ARAGUA 0.043 in MIRANDA and 1 in SANTIAGO DE CUBA. For the first time in Cuba a set of biochemical techniques was available for detecting the resistance mechanisms of Aedes aegypti which made it possible to create sound foundations for the easy rapid detection of the resistance of this species the main dengue vector in the Americas.

[Resistance to insecticides in Blattella germanica species strains from Santiago de Cuba]. / Resistencia a insecticidas en cepas de terreno de la especie Blattella germanica procedentes de Santiago de Cuba.

A study was conducted on the level of resistance to seven insecticides, namely, 3 organophosphate compounds (malathion, chlorpyrifos and pirimiphos-methyl), one carbamate (propoxur) and 1 pyrethroid (cypermethrin, deltamethrin and lambdacialotrine) of three field-collected strains of Blattella germanica (Linnaeus, 1767) from Santiago de Cuba. These strains showed high resistance levels to malathion, cypermethrin, deltamethrin and lambdacialotrine and low resistance to pirimiphos-methyl, and also they were susceptible to chlorpyrifos and propoxur. The levels of resistance to tested organophosphate insecticides such as malathion and pirimiphos-methyl and to pyrethroid compounds like cypermethrin, deltamethrin and lambdacialotrine may be related to the increased production of esterases as a mechanism of resistance. The value of frequency of the resistant genes for enzyme acetylcholinesterase was very low, therefore, the modified acetylcholinesterase is not involved in resistance to insecticides tested in the studied strains from Santiago de Cuba.