Pathogens Detection in the Small Hive Beetle (Aethina tumida (Coleoptera: Nitidulidae)).

Aethina tumida Murray is currently a worldwide emergent pest of Apis mellifera L. hives. Although the damaging effect on the colony stores and brood is well known, the possible role of these beetles as a disease carrier is not clear. This is the first report of DNA presence of the trypanosome honeybee parasite Lotmaria passim and Crithidia bombi, and the Apis mellifera filamentous virus (AmFV) in A. tumida. Further studies will be needed to determine if A. tumida is indeed a mechanical or biological vector of these pathogens.

Deformed wing virus type a and b in managed honeybee colonies of Argentina.

Apis mellifera is infected by more than 24 virus species worldwide, mainly positive-sense, single-stranded RNA viruses of the Dicistroviridae and Iflaviridae families. Among the viruses that infect honey bees, Deformed wing virus is the most prevalent and is present as three master variants DWV-A, B, and C. Given that the ectoparasitic mite Varroa destructor vectors these virus variants, recombination events between them are expected, and variants and their recombinants can co-exist in mites and honeybees at the same time. In this study, we detect, through RT-qPCR, the presence of DWV-A and B in the same samples of adult bees from colonies of Argentina. Total RNA was extracted from pools of ten adult bees from 45 apiaries distributed across the main beekeeping Provinces of Argentina (Buenos Aires, Santa Fe, Córdoba, Santiago del Estero, Río Negro, and Mendoza); then RT-qPCR reactions were performed to detect DWV-A and B, with specific primer pairs. After the amplifications, PCR products (204 and 660 bp amplicons for DWV-B, and ~250 bp for DWV-A) were purified and sequenced to verify that they corresponded to reported sequences, analyzing them using the Blast software. Of the 45 samples analyzed by RT-qPCR, over 90% were infected with DWV-A and 47% were also positive for DWV-B, where it was found in high prevalence specifically in colonies of A. mellifera of the Buenos Aires Province. Future studies will determine the impact of this type of the virus and its ability to recombine with the other DWV types in the apiaries of our country.

Heat shock proteins in Varroa destructor exposed to heat stress and in-hive acaricides.

Varroa destructor is one of the major pests that affect honeybees around the world. Chemical treatments are common to control varroosis, but mites possess biochemical adaptive mechanisms to resist these treatments, enabling them to survive. So far, no information is available regarding whether these pesticides can induce the expression of heat shock protein (Hsp) as a common protective mechanism against tissue damage. The aims of this study were to determine differences in heat shock tolerance between mites collected from brood combs and phoretic ones, and to examine patterns of protein expression of Hsp70 that occur in various populations of V. destructor after exposure to acaricides commonly employed in beekeeping, such as flumethrin, tau-fluvalinate and coumaphos. Curiously, mites obtained from brood cells were alive at 40 °C, unlike phoretic mites that reached 100% mortality, demonstrating differential thermo-tolerance. Heat treatment induced Hsp70 in mites 4 × more than in control mites and no differences in response were observed in phoretic versus cell-brood-obtained mites. Dose-response assays were carried out at increasing acaricide concentrations. Each population showed a different stress response to acaricides despite belonging to the same geographic region. In one of them, coumaphos acted as a hormetic stressor. Pyrethroids also induced Hsp70, but mite population seemed sensitive to this treatment. We concluded that Hsp70 could represent a robust biomarker for measuring exposure of V. destructor to thermal and chemical stress, depending on the acaricide class and interpopulation variability. This is relevant because it is the first time that stress response is analyzed in this biological model, providing new insight in host-parasite-xenobiotic interaction.

Lactobacillus kunkeei strains decreased the infection by honey bee pathogens Paenibacillus larvae and Nosema ceranae.

Due to their social behaviour, honey bees can be infected by a wide range of pathogens including the microsporidia Nosema ceranae and the bacteria Paenibacillus larvae. The use of probiotics as food additives for the control or prevention of infectious diseases is a widely used approach to improve human and animal health. In this work, we generated a mixture of four Lactobacillus kunkeei strains isolated from the gut microbial community of bees, and evaluated its potential beneficial effect on larvae and adult bees. Its administration in controlled laboratory models was safe for larvae and bees; it did not affect the expression of immune-related genes and it was able to decrease the mortality associated to P. larvae infection in larvae and the counts of N. ceranae spores from adult honey bees. These promising results suggest that this beneficial microorganism's mixture may be an attractive strategy to improve bee health. Field studies are being carried out to evaluate its effect in naturally infected colonies.

Efficacy of Essential Oils of Thymus vulgaris and Origanum vulgare on Echinococcus granulosus.

The aim of the present work was to determine the in vitro effect of T. vulgaris and O. vulgare essential oils against E. granulosus protoscoleces and cysts. Essential oils were added to the medium resulting in thymol final concentrations of 10 µg/mL. The essential oils had a time-dependent effect provoking the complete loss of protoscolex viability after 72 days of postincubation. The results were confirmed at the ultrastructure level. Loss of infectivity in protoscoleces incubated with O. vulgare after 60 days was observed. On the other hand, the weight of cysts recorded in mice inoculated with T. vulgaris treated protoscoleces was significantly lower than that obtained in control group. Gamma-glutamyl-transpeptidase activity was readily detected in the culture supernatant of protoscoleces treated either with the essential oils or thymol. T. vulgaris and O. vulgare essential oils and thymol can induce cell apoptosis of protoscoleces after short incubation times. The efficacy of T. vulgaris and O. vulgare essential oils was also demonstrated in vitro on E. granulosus murine cysts. Our data suggest that essential oils of T. vulgaris and O. vulgare have anthelmintic effect against protoscoleces and cysts of E. granulosus.

Synergistic effect of surfactin from Bacillus subtilis C4 and Achyrocline satureioides extracts on the viability of Paenibacillus larvae.

The aim of this work was to determine the in vitro effect of the mixture between the lipopeptide surfactin, synthesized by Bacillus subtilis C4 (strain isolated from honey) and the most active vegetal extract from Achyrocline satureioides, a traditional medicinal plant, on local strains of Paenibacillus larvae, the agent of American Foulbrood in honeybees. Five P. larvae strains isolated in Córdoba, Argentina, were phenotypically characterized. These and 12 other P. larvae strains from different regions of Argentina were analysed. The antimicrobial activities of the essential oil, hexane (HE) and benzene extracts from A. satureioides were assessed against P. larvae and the HE showed the highest anti-P. larvae activity. A combination of the biosurfactant surfactin, produced by B. subtilis C4, and the HE of A. satureioides revealed a synergistic action on P. larvae. The effective surfactin concentration in the mixture decreased from 32 to 1 µg ml(-1) and the HE concentration from 32 to 4 µg ml(-1), values similar or equal to minimal inhibitory concentrations observed for oxytetracycline. The fractional inhibitory concentration index confirmed synergism in 4 strains and partial synergism in one strain. The combination of surfactin synthesized by B. subtilis C4 and the HE from A. satureioides could be a natural alternative to help beekeepers to combat the American foulbrood agent P. larvae.

Genetic structure of Varroa destructor populations infesting Apis mellifera colonies in Argentina.

Although mitochondrial DNA mapping of Varroa destructor revealed the presence of several haplotypes, only two of them (Korean and Japanese haplotypes) were capable to infest Apis mellifera populations. Even though the Korean haplotype is the only one that has been reported in Argentina, these conclusions were based on mites sampled in apiaries from a specific geographical place (Buenos Aires province). To study mites from several sites of Argentina could reveal the presence of the Japanese genotype, especially considering sites near to Brazil, where Japanese haplotype was already detected. The aim of this work was to study the genetic structure of V. destructor populations from apiaries located in various provinces of Argentina, in order to determine the presence of different haplotypes. The study was carried out between January 2006 and December 2009. Phoretic adult Varroa mites were collected from honey bee workers sampled from colonies of A. mellifera located in Entre Ríos, Buenos Aires, Corrientes, Río Negro, Santa Cruz and Neuquén provinces. Twenty female mites from each sampling site were used to carry out the genetic analysis. For DNA extraction a nondestructive method was used. DNA sequences were compared to Korean haplotype (AF106899) and Japanese haplotype (AF106897). All DNA sequences obtained from mite populations sampled in Argentina, share 98% of similitude with Korean Haplotype (AF106899). Taking into account these results, we are able to conclude that Korean haplotype is cosmopolite in Argentina.

Bioactivity of Rosmarinus officinalis essential oils against Apis mellifera, Varroa destructor and Paenibacillus larvae related to the drying treatment of the plant material.

In this study, chemical composition, physicochemical properties and bioactivity of two essential oils of Rosmarinus officinalis extracted from plant material with different drying treatments against Apis mellifera, Varroa destructor and Paenibacillus larvae were assessed. The lethal concentration 50 (LC50) for mites and bees was estimated using a complete exposure method test. The broth microdilution method was followed in order to determine the minimum inhibitory concentrations (MICs) of the essential oils against P. larvae. Physicochemical properties were similar in both the essential oils, but the percentage of components showed certain differences according to their drying treatment. ß-Myrcene and 1,8-cineole were the main constituents in the oils. The LC50 for complete exposure method at 24, 48 and 72 h was minor for mites exposed to R. officinalis essential oil dried in oven conditions. MIC values were 700-800 µg mL(-1) and 1200 µg mL(-1) for R. officinalis dried in air and oven conditions, respectively. The results reported in this research show that oil toxicity against V. destructor and P. larvae differed depending on the drying treatment of the plant material before the distillation of essential oil.

Evaluation of culture media for Paenibacillus larvae applied to studies of antimicrobial activity.

This study was conducted to compare different liquid culture media for Paenibacillus larvae growth in order to find the best one to be used in studies on activity of antimicrobial substances, such as essential oils. P. larvae presented poor growth in usual broths such as Mueller-Hinton, commonly employed in antimicrobial activity assays. Growth in liquid media was evaluated using Paenibacillus larvae strains isolated from hives located in different geographical zones. The MYT medium (Mueller-Hinton broth, yeast extract and thiamine) was selected out of the eight liquid media analyzed, as it proved to be the most adequate due to its higher absorbance at 620 nm. The following mean values were obtained from the four P. larvae strains: 0.227 +/- 0.016 for the Cobo strain, 0.279 +/- 0.015 for La Plata strain, 0.758 +/- 0.020 for Mechongué strain and 0.244 +/- 0.0079 for Sierra de los Padres strain, respectively.

Evaluation of culture media for Paenibacillus larvae applied to studies of antimicrobial activity / Evaluación de medios de cultivo para el crecimiento de Paenibacillus larvae aplicables en estudios de actividad antimicrobiana

This study was conducted to compare different liquid culture media for Paenibacillus larvae growth in order to find the best one to be used in studies on activity of antimicrobial substances, such as essential oils. P. larvae presented poor growth in usual broths such as Mueller-Hinton, commonly employed in antimicrobial activity assays. Growth in liquid media was evaluated using Paenibacillus larvae strains isolated from hives located in different geographical zones. The MYT medium (Mueller-Hinton broth, yeast extract and thiamine) was selected out of the eight liquid media analyzed, as it proved to be the most adequate due to its higher absorbance at 620 nm. The following mean values were obtained from the four P. larvae strains: 0.227 ± 0.016 for the Cobo strain, 0.279 ± 0.015 for La Plata strain, 0.758 ± 0.020 for Mechongué strain and 0.244 ± 0.0079 for Sierra de los Padres strain, respectively.

Este trabajo está orientado a comparar diferentes medios de cultivo líquidos para el crecimiento de Paenibacillus larvae. El objetivo fue encontrar el más apropiado para utilizar en estudios de actividad antimicrobiana de diferentes sustancias, tales como aceites esenciales. P. larvae presenta un crecimiento débil en medios de cultivo como el Mueller-Hinton, comúnmente usado en ensayos de actividad antimicrobiana. Se evaluó el crecimiento en caldos de cultivo de cepas aisladas de colmenas ubicadas en diferentes zonas geográficas. De los ocho medios analizados, el MYT (Mueller-Hinton, extracto de levadura y tiamina) mostró ser el más apropiado, en éste se observó el mayor valor de absorbancia a 620 nm. Los valores obtenidos en promedio para los cuatro aislamientos de P. larvae evaluados fueron 0,227 ± 0,016 (cepa de Cobo); 0,279 ± 0,015 (cepa de La Plata); 0,758 ± 0,020 (cepa de Mechongué) y 0,244 ± 0,0079 (cepa de Sierra de los Padres).

[Inhibition of Paenibacillus larvae employing a mixture of essential oils and thymol]. / Inhibición de Paenibacillus larvae empleando una mezcla de aceites esenciales y timol.

In vitro antimicrobial activity of a mixture of two essential oils and thymol against Paenibacillus larvae, causal agent of American Foulbrood (AFB), was evaluated. The essential oils were extracted from cinnamon (Cinnamomum zeylanicum) and thyme (Thymus vulgaris). The third component used, thymol, is the major component of the essential oil of thyme which contains 39.9% of thymol. Minimal inhibitory concentration (MIC) in Mueller-Hinton broth by the tube dilution method and minimal bactericide concentration (MBC) on MYPGP agar were evaluated. Thyme registered MIC values of 150-250 microg/ml and MBC values of 200-300 microg/ml, while the MIC and MBC values obtained for cinnamon were of 50-100 microg/ml and 100-125 microg/ml. Thymol showed similar MIC and MBC values of 100-150 microg/ml. No significant differences between the bacterial strains were detected, but significant differences between essential oils and thymol activity were registered (P<0,01). An inhibitory synergetic effect on AFB was observed reducing MIC and MBC values due to the use of a mixture of 62.5% of thyme, 12.5% of cinnamon and 25% of thymol.

A new product with formic acid for Varroa jacobsoni Oud. control in Argentina. I. Efficacy.

An organic product based on formic acid in a gel matrix was evaluated for use in Varroa control under autumnal climatic conditions in Argentina. Twenty colonies each received two gel packets with formic acid in two applications and numbers of falling mites were registered. After this treatment colonies received two other acaricides in order to compare efficacy. Average final efficacy in colonies treated with the organic product was 92% with a low variability. The gel matrix kept an adequate formic acid concentration inside the colonies with only two applications. This product is, therefore, a good alternative for Varroa control because it is organic, easy to use and presents a low variability in final efficacy between colonies. No queen, brood, or adult honeybee mortality was registered.

[Pericarditis secondary to the rupture of a hydatid cyst]. / Pericarditis secundaria a rotura de un quiste hidatídico.

Cardiac hydatic cysts are rare and represent 0.5 to 2% of all hydatic cysts in humans, but usually associated with fatal complications. We report a case of a male 27 years old with a hydatid cyst located in left ventricle asymptomatic until rupture. It was diagnosed by two-dimensional echocardiogram in a control.

[Myocardial revascularization in the female sex: increased surgical risk or referral bias?]. / Revascularización miocárdica en el sexo femenino: mayor riesgo quirúrgico o sesgo de referencia?

During the last fifteen years the increased morbimortality of coronary surgery in women has been widely documented. Traditionally, the increased risk has been attributed to the smaller size of the coronary arteries of female patients. However, there are data in our series and in various others that lead to suspicion that there might exist a referral bias, which causes women to be referred to a surgeon at a more advanced age and in a worse clinical condition than men. This bias, if it is present, could contribute to the worse results obtained in coronary artery bypass surgery in women. The only way to determine if this is valid is by applying similar criteria of management to both sexes. Only in this way will it be possible to ascertain if the high morbimortality of myocardial revascularization surgery in women is due to factors that are related to the gender itself or to referral bias stated above.

Contrapulsación intraaórtica en cirugía coronaria: factores predictivos de mortalidad hospitalaria / Intraaortic countenpulsation in coronary surgery: prediction factors of hospital mortality

La contrapulsación aórtica (CPIA) es el método más básico y difundido de asistencia circulatoria. Su utilidad en el síndrome de bajo gasto postcardiotomía aún comporta alta mortalidad. Pretendemos identiificar factores relacionados con el perfil preoperatoriodel paciente y el procedimiento realizado, que tenga carácter predictivo de mortalidad hospitalaria (MH) en la utilización de CPIA por bajo gasto tras cirugía coronaria. Entre septiembre de 1995 y abril de 1998 hemos intervenido 382 pacientes coronarios. Se utilizó CPIA en 54 casos (14 por ciento). Analizamos 50 variables pre e intraoperatorias de potencial carácter predictivo en el grupo A (29 supervivientes) y B (25 fallecidos). La MH por CPIA en coronarios fue del 46 por ciento. Todos los scores de riesgo preoperatorio fueron más elevados en el grupo B, aunque el score de Parsonnet mostró diferencias significativas (grupo A: 6 vs B: 12,9) (p<0,01). La mayor utilización de venas (grupo A:1,3 vs B:2) (p<0,05), el menor uso de injertos arteriales (A:79 por ciento vs B:56 por ciento) y el carácter emergente de la cirugía (A:7 por ciento vs B:20 por ciento) también se relacionaron con la MH. La CPIA en el sindrome postcardiotomía tras revascularización coronaria aún presenta altas cifras de MH. En nuestra experiencia la MH se asocia a un perfil de riesgo elevado del paciente, así como con el tipo de injerto empleado y con el carácter emergente de la cirugía

Contrapulsación intraaórtica en cirugía coronaria: factores predictivos de mortalidad hospitalaria / Intraaortic countenpulsation in coronary surgery: prediction factors of hospital mortality

La contrapulsación aórtica (CPIA) es el método más básico y difundido de asistencia circulatoria. Su utilidad en el síndrome de bajo gasto postcardiotomía aún comporta alta mortalidad. Pretendemos identiificar factores relacionados con el perfil preoperatoriodel paciente y el procedimiento realizado, que tenga carácter predictivo de mortalidad hospitalaria (MH) en la utilización de CPIA por bajo gasto tras cirugía coronaria. Entre septiembre de 1995 y abril de 1998 hemos intervenido 382 pacientes coronarios. Se utilizó CPIA en 54 casos (14 por ciento). Analizamos 50 variables pre e intraoperatorias de potencial carácter predictivo en el grupo A (29 supervivientes) y B (25 fallecidos). La MH por CPIA en coronarios fue del 46 por ciento. Todos los scores de riesgo preoperatorio fueron más elevados en el grupo B, aunque el score de Parsonnet mostró diferencias significativas (grupo A: 6 vs B: 12,9) (p<0,01). La mayor utilización de venas (grupo A:1,3 vs B:2) (p<0,05), el menor uso de injertos arteriales (A:79 por ciento vs B:56 por ciento) y el carácter emergente de la cirugía (A:7 por ciento vs B:20 por ciento) también se relacionaron con la MH. La CPIA en el sindrome postcardiotomía tras revascularización coronaria aún presenta altas cifras de MH. En nuestra experiencia la MH se asocia a un perfil de riesgo elevado del paciente, así como con el tipo de injerto empleado y con el carácter emergente de la cirugía (AU)

[Aortic intramural hematoma. An atypical pattern equivalent to aortic dissection]. / Hematoma intramural aórtico. Un patrón atípico equivalente a la disección aórtica.

A case is presented of a hypertensive woman who had suffered a stabbing back pain for some three hours, with mild irradiation to precordium and accompanied by vegetative signs. A sinusal rhythm and negative T waves of little depth were seen on the ECG. A transthoracic bidimensional echocardiogram (TTE) showed a normal left ventricle with a somewhat dilated aortic root and the existence of a double echo running parallel to the anterior wall of the aorta but non-ondulating and without a visible intimal flap. Because of suspected aortic dissection an urgent contrasted CAT and a transesophageal echocardiogram were performed. These were informed as an aneurysm of the aortic root with mural thrombus from the ascending to descending aorta, but with no existing intimal flap suggesting dissection. A cardiac catheterization showed a mildly some dilated aortic root without dissection signs and normal left ventricle and coronary arteries. The patient was presented for surgical evaluation but, since no dissection was present, was not considered urgent surgery; she was admitted to the coronary unit and died 48 hours later in a situation of acute pericardial tamponade, documented by TTE, surely due to rupture of the aortic root to pericardial sack. This way of presenting threatened aorta rupture that has been only recently recognized is discussed, as well as some misconceptions which must be avoided.

Comparative evaluation of pentoxifylline, buflomedil, and nifedipine in the treatment of intermittent claudication of the lower limbs.

In the light of the current controversy surrounding the use of hemorheologic and vasodilator drugs in the treatment of peripheral arteriosclerosis, a comparative study was designed in order to evaluate the efficacy of pentoxifylline, buflomedil, and nifedipine in 45 patients with peripheral arterial disease (Fontaine stage II). The patients in this prospective randomized study were divided into three groups: 15 patients received pentoxifylline treatment (1,200 mg/day), 15 were treated with buflomedil (600 mg/day), and 15 with nifedipine (60 mg/day). Response to treatment was assessed at the start of the study and after forty-five and ninety days, by clinical examination, Doppler test, strain test, and digital occlusion plethysmography using a strain gauge ring. Pentoxifylline was significantly more effective (P < 0.05) than buflomedil and nifedipine at ninety days in improving walking performance, resting toe pressure, resting and postexercise ankle/brachial pressure ratio, and basal/postischemic toe-pulse ratio. Significant differences within groups were also noted for initial claudication, toe peak-flow time, pulse reappearance time (PRT/2), and maximum postischemic flow time, together with significant intergroup variables. In conclusion, pentoxifylline proved more effective than the other drugs tested in: 1. improving distal pressure and resting microcirculatory blood flow; 2. increasing postexercise distal flow, ratios, and pressures and enabling faster recuperation of basal pulse rates; 3. increasing initial claudication distance in the strain test within the test group and achieving a greater absolute subjective claudication distance than that obtained using the other treatments.

Early open mitral commissurotomy: long-term results.

It has frequently been suggested that early mitral commissurotomy could improve long-term results in patients with severe mitral stenosis. However, the real advantages of this procedure have yet to be demonstrated. To evaluate this hypothesis, we retrospectively studied 397 patients who underwent operation for mitral stenosis in our unit between 1978 and 1988. Forty of these patients (group I) fulfilled the criteria for early mitral commissurotomy: being young (average age 33 years), being asymptomatic or showing few symptoms, and being in sinus rhythm. The remaining 357 patients who underwent operation during the same period of time served as control (group II). Mitral valve replacement (p < 0.05) and associated tricuspid annuloplasty (p < 0.05) occurred less frequently in patients from group I than it did in patients from group II. Survival after 11 years for patients with early mitral commissurotomy was 100%, 90% of whom were in functional class I, were in sinus rhythm, and were receiving no medication whatsoever. Ninety-six percent of these patients were free of complications after 11 years, compared with 73% of patients in group II (p < 0.05). In our opinion, these results support the use of early mitral commissurotomy for young patients in sinus rhythm, who are symptom free, who are and who have a mitral valve area is 1.3 cm2 or less.

Pulmonary valve endocarditis during puerperal sepsis.

A case of pulmonary valve endocarditis caused by Staphylococcus aureus during puerperal sepsis in a female patient is reported. The M-mode and two dimensional echocardiographic finding are described. A review of the literature shows that this entity is rare. A large vegetation in the leaflet of the pulmonary valve was excised and the patient recovered after a full course of antibiotics.