Disintegrins extracted from totonacan rattlesnake (Crotalus totonacus) venom and their anti-adhesive and anti-migration effects on MDA-MB-231 and HMEC-1 cells.

Disintegrins are low molecular weight cysteine-rich proteins (4-14 kDa) that are isolated mainly from viperid snake venom. Due to their potential as lead compounds for binding and blocking integrin receptors, snake venom disintegrins have become one of the most studied venom protein families. The aim of this study was to obtain disintegrins from C. totonacus venom and evaluate their capability to bind and block integrin receptors. The C. totonacus disintegrin fraction (totonacin) represents two disintegrin isoforms obtained from C. totonacus venom. These disintegrins showed extracellular-matrix (ECM) protein adhesion and migration inhibitory effects on MDA-MB-231 and HMEC-1 cells. Totonacin (3 µM) inhibited MDA-MB-231 cell adhesion to the ECM proteins, fibronectin, vitronectin, and laminin by 31.2, 44.0, and 32.1, respectively. Adhesion inhibition to fibronectin, vitronectin, and laminin observed on HMEC-1 cells was 42.8, 60.8, and 51%, respectively. In addition, totonacin (3 µM) significantly inhibited MDA-MB-231 and HMEC-1 cell migration (41.4 and 48.3%, respectively). Totonacin showed more potent cell adhesion inhibitory activity toward vitronectin in both cell lines. These results suggest a major affinity of totonacin toward αVß3, α8ß1, αVß5, αVß1, and αIIbß3 integrins. In addition, the inhibitory effect observed on MDA-MB-231 and HMEC-1 cell migration reinforces the evidence of an interaction between these disintegrins and αVß3 integrin, which plays a key role in migration and angiogenesis.

Invasive pneumococcal disease in hospitalised children from Lima, Peru before and after introduction of the 7-valent conjugated vaccine.

The objective of this study was to determine the serotype distribution and antibiotic resistance of invasive pneumococcal disease (IPD) strains in children from Lima, Peru, before and after the introduction of the 7-valent pneumococcal conjugate vaccine (PCV7), which was introduced in the national immunisation program on 2009. We conducted a prospective, multicentre, passive surveillance IPD study during 2006-2008 and 2009-2011, before and right after the introduction of PCV7 in Peru. The study was performed in 11 hospitals and five private laboratories in Lima, Peru, in patients <18 years old, with sterile site cultures yielding Streptococcus pneumoniae. In total 159 S. pneumoniae isolates were recovered. There was a decrease in the incidence of IPD in children <2 years old after the introduction of PCV7 (18.4/100 000 vs. 5.1/100 000, P = 0.004). Meningitis cases decreased significantly in the second period (P = 0.036) as well as the overall case fatality rate (P = 0.025), including a decreased case fatality rate of pneumonia (16.3% to 0%, P = 0.04). PCV7 serotypes showed a downward trend. Vaccine-preventable serotypes caused 78.9% of IPD cases, mainly 14, 6B, 5, 19F and 23F. A non-significant increase in erythromycin resistance was reported. Our findings suggest that the introduction of PCV7 led to a significant decrease of IPD in children under 2 years old and in the overall case fatality rate.

Eight-plate epiphysiodesis: are we creating an intra-articular deformity?

Aims: Guided growth using eight-plates is commonly used for correction of angular limb deformities in growing children. The principle is of tethering at the physeal periphery while enabling growth in the rest of the physis. The method is also applied for epiphysiodesis to correct limb-length discrepancy (LLD). Concerns have been raised regarding the potential of this method to create an epiphyseal deformity. However, this has not been investigated. The purpose of this study was to detect and quantify the occurrence of deformities in the proximal tibial epiphysis following treatment with eight-plates. Patients and Methods: A retrospective study was performed including 42 children at a mean age of 10.8 years (3.7 to 15.7) undergoing eight-plate insertion in the proximal tibia for correction of coronal plane deformities or LLD between 2007 and 2015. A total of 64 plates were inserted; 48 plates (34 patients) were inserted to correct angular deformities and 16 plates (8 patients) for LLD. Medical records, Picture Archive and Communication System images, and conventional radiographs were reviewed. Measurements included interscrew angle, lateral and medial plateau slope angles measured between the plateau surface and the line between the ends of the physis, and tibial plateau roof angle defined as 180° minus the sum of both plateau angles. Measurements were compared between radiographs performed adjacent to surgery and those at latest follow-up, and between operated and non-operated plateaus. Statistical analysis was performed using BMDP Statistical Software. Results: Slope angle increased in 31 (49.2%) of operated epiphyses by a mean of 5° (1° to 23°) compared with 29 (31.9%) in non-operated epiphyses (p = 0.043). Roof angle decreased in 29 (46.0%) of operated tibias and in 25 (27.5%) of non-operated ones by a mean of 5° (1° to 18°) (p = 0.028). Slope angle change frequency was similar in patients with LLD, varus and valgus correction (p = 0.37) but roof angle changes were slightly more frequent in LLD (p = 0.059) and correlated with the change in inter screw angles (r = 0.74, p = 0.001). Conclusion: The use of eight-plates in the proximal tibia for deformity correction and limb-length equalization causes a change in the bony morphology of the tibial plateau in a significant number of patients and the effect is more pronounced in the correction of LLD. Cite this article: Bone Joint J 2018;100-B:1112-16.

Shiga toxin-producing Escherichia coli (STEC) O22:H8 isolated from cattle reduces E. coli O157:H7 adherence in vitro and in vivo.

PROBLEM ADDRESSED: Shiga toxin-producing Escherichia coli (STEC) are a group of bacteria responsible for food-associated diseases. Clinical features include a wide range of symptoms such as diarrhea, hemorrhagic colitis and the hemolytic uremic syndrome (HUS), a life-threatening condition. OBJECTIVE: Our group has observed that animals naturally colonized with STEC strains of unknown serotype were not efficiently colonized with E. coli O157:H7 after experimental infection. In order to assess the basis of the interference, three STEC strains were isolated from STEC persistently-colonized healthy cattle from a dairy farm in Buenos Aires, Argentina. METHODS AND RESULTS: The three isolated strains are E. coli O22:H8 and carry the stx1 and stx2d genes. The activatable activity of Stx2d was demonstrated in vitro. The three strains carry the adhesins iha, ehaA and lpfO113. E. coli O22:H8 formed stronger biofilms in abiotic surface than E. coli O157:H7 (eae+, stx2+) and displayed a more adherent phenotype in vitro towards HeLa cells. Furthermore, when both serotypes were cultured together O22:H8 could reduce O157:H7 adherence in vitro. When calves were intragastrically pre-challenged with 108 CFU of a mixture of the three STEC strains and two days later challenged with the same dose of the strain E. coli O157:H7 438/99, the shedding of the pathogen was significantly reduced. CONCLUSIONS: These results suggest that E. coli O22:H8, a serotype rarely associated with human illness, might compete with O157:H7 at the bovine recto-anal junction, making non-O157 carrying-calves less susceptible to O157:H7 colonization and shedding of the bacteria to the environment.

Lactoferrin concentration in breast milk of mothers of low-birth-weight newborns.

OBJECTIVES: Lactoferrin (LF) is a breast milk glycoprotein with protective effects against neonatal infections, mainly in premature and low-birth-weight (LBW) neonates. The aims of this study were to determine LF concentration in breast milk of mothers of LBW infants during the first 2 months postpartum, and to identify the factors associated with LF concentration. STUDY DESIGN: Prospective study conducted as a part of an ongoing clinical trial in three Neonatal Units in Peru. We included 346 mothers of neonates with a birth weight <2000 g. We measured LF concentration in four stages of lactation using a commercial enzyme-linked immunosorbent assay kit. Multivariate analysis was performed to assess the association between maternal and neonatal factors, and LF concentration. RESULTS: We collected 695 milk samples. LF mean concentration±standard deviation was 14.92±7.96 mg ml-1 in colostrum (n=277), 10.73±5.67 in transitional milk (n=55), 10.34±6.27 at 1 month (n=259) and 8.52±6.47 at 2 months (n=104). There was a significant difference in LF concentration between different stages of lactation (P<0.001). Mothers with higher LF concentration in colostrum had higher values in the following 2 months. High maternal income and multiple gestation were significantly associated with higher LF levels; in contrast, maternal peripartum infections and male neonatal gender were associated with lower LF levels. CONCLUSIONS: LF concentration in breast milk of mothers of LBW infants was high and remained elevated even at 1 and 2 months postpartum. LF concentration in colostrum was higher in mothers with higher income and multiple pregnancies, and lower in mothers with peripartum infections.

Evaluation of Culture Conditions to Obtain Fatty Acids from Saline Microalgae Species: Dunaliella salina, Sinecosyfis sp., and Chroomonas sp.

The use of the saline microalgae, Dunaliella salina, Sinecosyfis sp., and Chroomonas sp., was explored as an alternative source for the production of fatty acids using fertilizer and glycerol as culture media. The nutrient medium used contained "Nutrifoliar," a commercial fertilizer, and/or glycerol, in natural sea water. The microalgae were placed in cultures with different conditions. The parameters that favored the largest production of fatty acids were 24 hours of agitation and illumination, 1620 L/day of air supply, 2.25 L of air/min, and a temperature of 32°C using "Nutrifoliar" as the culture media. Results indicated that, from 3 g of microalgae in wet base of Chroomonas sp., 54.43 mg of oil was produced. The chromatographic characterization of oil obtained revealed the presence of essential fatty acids such as 9,12,15-octadecatrienoic acid (omega-3) and 4,7,10-hexadecatrienoic acid (omega-6) from the species Dunaliella salina. On the other hand, 9,12-octadecadienoic acid (omega-6) and cis-11-eicosenoic acid (omega-9) were identified from the species Chroomonas sp. The temperature variations played an important role in the velocity of growth or the production of the algae biomass, the amount of oil, and the ability to produce fatty acids.

The intranasal vaccination of pregnant dams with Intimin and EspB confers protection in neonatal mice from Escherichia coli (EHEC) O157:H7 infection.

Enterohemorrhagic Escherichia coli (EHEC) O157:H7 is responsible for intestinal disease and hemolytic uremic syndrome (HUS), a serious systemic complication which particularly affects children. In this study, we evaluated whether passive immunization protects from EHEC O157:H7 colonization and renal damage, by using a weaned BALB/c mouse model of infection. Recombinant proteins EspB and the carboxyl-terminal fragment of 280 amino acids of γ-intimin (γ-IntC280) were used in combination with a macrophage-activating lipopeptide-2 (MALP) adjuvant to immunize pregnant mice by the intranasal route. Neonatal mice were allowed to suckle vaccinated or sham-vaccinated dams until weaning when they were challenged by the oral route with a suspension of an E. coli O157:H7 Stx2+ strain. The excretion of the inoculated strain was followed for 72h. All vaccinated dams exhibited elevated serum IgG response against both γ-Int C280 and EspB. Passive immunization of newborn mice resulted in a significant increase in serum IgG titers against γ-Int C280 and a slight increase in EspB-specific antibodies. The neonates from vaccinated dams showed a significant reduction in EHEC O157:H7 colonization 48h post challenge. In addition, the level of plasma urea concentration, a marker of renal failure, was significantly higher in offsprings of sham-vaccinated mice. In conclusion, vaccination of pregnant dams with γ-Int C280 and EspB could reduce colonization and systemic toxicity of EHEC O157:H7 in their suckling offsprings.

Gastric mucosa injury quantification in an ischemia - Reperfusion experimental model.

Gastric ischemia - reperfusion (I/R) injury is an important clinical problem, which is developed in more than 80% of critically ill patients. I/R is caused by interruption of blood supply to an organ or tissue followed by blood reflow into the exposed area, leading to multiple organ failure and death. Gastric reactance has been proposed to measure tissue injury caused by ischemia. The present study evaluates a new method to quantify gastric tissue damage due to I/R, and assess its relation to gastric reactance changes. Twenty Wistar rats were randomly assigned to 4 groups: control, ischemia, I/R 30 min, I/R 1 h. Local gastric ischemia was induced by clamping the celiac artery for 30 min and reperfusion was done for 30-60 min. In all groups, gastric impedance was measured, and then gastric mucosa samples were taken for light microscopy. There were statistical significant differences (p <;0.05) among the groups with respect to the index of gastric injury proposed, which was greater in I/R 1 h group. Also, impedance parameters increased in I/R groups with respect to control, and ischemia groups. The proposed index of gastric injury allowed gastric mucosa damage quantification, and it was related with gastric impedance increase, which is an objective method to evaluate tissue injury.

Impact of Infection Dose and Previous Serum Antibodies against the Locus of Enterocyte Effacement Proteins on Escherichia coli O157:H7 Shedding in Calves following Experimental Infection.

Escherichia coli O157:H7 is the main causative agent of haemolytic uremic syndrome. Cattle are the main reservoir of these bacteria, and have been shown to develop immune response to colonization. Our aim was to investigate the faecal shedding pattern of E. coli O157:H7 in calves challenged intragastrically with either 10(8) or 10(10) CFU, as well as the ability of specific preexisting antibodies to reduce shedding of the pathogen. Shedding was analysed by direct counting as well as enrichment of rectoanal mucosal swabs. Statistical analysis was performed using a linear model for repeated measures with and without the inclusion of preexisting antibodies against the carboxy-terminal fraction of intimin-γ (γ-intimin C280) as a covariable. Results suggest that there is a statistical difference in the area under the shedding curves between both doses for 14 as well as 28 days after challenge (p = 0.0069 and 0.0209, resp.). This difference is increased when the prechallenge antibodies are taken into account (p = 0.0056 and 0.0185). We concluded that the bacterial dose influences shedding on calves experimentally challenged and that preexisting antibodies against E. coli O157:H7 γ-intimin C280 could partially reduce faecal excretion.

Physiopathological effects of Escherichia coli O157:H7 inoculation in weaned calves fed with colostrum containing antibodies to EspB and Intimin.

Escherichia coli O157:H7 is responsible for severe intestinal disease and hemolytic uremic syndrome (HUS), a serious systemic complication which particularly affects children. Cattle are the primary reservoir for E. coli O157:H7 and the main source of infection for humans. In this study, we evaluated the ability of transferred maternal colostral antibodies against γ-Intimin C280 and EspB, to protect young weaned calves from E. coli O157:H7 infection. Hyperimmune colostra were obtained by immunization of pregnant cows with a mix of the mentioned antigens. All vaccinated cows mounted a significant IgG response against γ-Intimin C280, and EspB in sera and colostra. Colostrum-fed calves also exhibited high serum IgG titers against γ-Intimin C280 and EspB along with a rise in mucosal γ-Intimin C280-specific IgG antibodies at recto-anal junction and ileum. Additionally, 70 day-old calves received a challenge with E. coli O157:H7 but no reduction in total bacterial shedding or frequency of E. coli O157:H7 excretion from these calves was observed. Most tissue samples showed granulocyte focal infiltrations of the lamina propria and enterocyte erosion. In conclusion, up to the 70th day, the passively acquired γ-Intimin-C280 and EspB-IgG antibodies present in sera and recto-anal mucosa reached a titer insufficient to reduce EHEC O157 shedding and damages of experimentally inoculated young calves.

Addition of cholesterol-loaded cyclodextrins to the thawing extender: effects on boar sperm quality.

The aim of the present study was to evaluate the effect that the addition of cholesterol-loaded cyclodextrins (CLC) to the thawing extender has on the quality of frozen-thawed boar sperm. Pooled semen (n = 5) from three boars was used for the experiments. The semen was cryopreserved with an egg-yolk-based extender, it was diluted after thawing in Beltsville thawing solution (BTS) supplemented with different concentrations of CLC (0, 12.5, 25, 50 or 100 mg/500 × 10(6) sperm), and these samples were incubated at 37°C for 150 min. The following parameters of sperm quality were evaluated 30 and 150 min after incubation: sperm with intact plasma membrane (SIPM; %), sperm with normal acrosomal ridge (NAR; %), total motile sperm (TMS; %), progressively motile sperm (PMS; %) and kinetic parameters. Both SIPM and NAR increased (p < 0.05) when the thawing extender was supplemented with 12.5, 25 and 50 mg CLC/500 × 10(6) sperm. Nevertheless, motility decreased (p < 0.05) when the concentration of CLC exceeded 12.5 mg CLC/500 × 10(6) sperm. In conclusion, our results suggest that the supplementation of thawing extenders with CLC improves sperm viability and reduces acrosome damage after freezing/thawing.

Multiplex real-time PCR for detection of Campylobacter, Salmonella, and Shigella.

Infectious diarrhea can be classified based on its clinical presentation as noninflammatory or inflammatory disease. In developing countries, among inflammatory diarrhea cases, Shigella is the most common cause, followed by Campylobacter and Salmonella. Because the time frame in which treatment choices must be made is short and conventional stool cultures lack good sensitivity, there is a need for a rapid, sensitive, and inexpensive detection technique. The purpose of our study was to develop a multiplex real-time PCR procedure to simultaneously identify Campylobacter spp., Salmonella spp., and Shigella spp. Primers were designed to amplify the invA, ipaH, and 16S rRNA genes simultaneously in a single reaction to detect Salmonella, Shigella, and Campylobacter, respectively. Using this approach, we correctly identified 102 of 103 strains of the targeted enteropathogens and 34 of 34 other pathogens. The melting temperatures were 82.96 ± 0.05 °C for invA, 85.56 ± 0.28 °C for ipaH, and 89.21 ± 0.24 °C for 16S rRNA. The limit of accurate quantification for the assay in stool samples was 10(4) CFU g(-1); however, the limit of detection was 10(3) CFU g(-1). This assay is a simple, rapid, inexpensive, and reliable system for the practical detection of these three enteropathogens in clinical specimens.

Is sperm freezability related to the post-thaw lipid peroxidation and the formation of reactive oxygen species in boars?

The aim of the present study was to determine whether the levels of reactive oxygen species (ROS) substances production and the levels of lipid peroxidation of the sperm membrane were related to the quality that the ejaculates exhibited after cryopreservation in boars. Ejaculates from 42 healthy boars were used in this study and they were cryopreserved with the lactose-egg yolk extender (LEY). Several sperm quality parameters were assessed by flow cytometry in samples incubated for 30 and 150 min at 37 °C after thawing: the percentage of sperm with intact plasma membrane (SIPM), intracellular reactive oxygen substances production through mean of DCF fluorescence intensity of total sperm (mean-DCF) and the percentage of viable and non-viable sperm containing oxidized BODIPY (VSOB and NVSOB). In addition, the percentages of total motile (TMS) and progressively motile sperm (PMS) were assessed at the same incubation times with a computer-assisted sperm analysis system. The classification of the ejaculates into good or bad freezers was performed through hierarchical cluster analysis from SIPM and TMS at 150 min post-thawing. The ejaculates of those males classified as good freezers exhibited higher (p < 0.05) SPIM, TMS and PMS than the bad freezers, although both groups presented similar (p > 0.05) VSOB, NVSOB and mean-DCF. Therefore, these results show that lipid peroxidation and the amount of reactive oxygen substances in the sperm after cryopreservation are similar between boars classified as good or bad freezers.

Vaccination of pregnant cows with EspA, EspB, γ-intimin, and Shiga toxin 2 proteins from Escherichia coli O157:H7 induces high levels of specific colostral antibodies that are transferred to newborn calves.

Enterohemorrhagic Escherichia coli (EHEC) O157:H7 is a major cause of intestinal disease and hemolytic uremic syndrome, a serious systemic complication that particularly affects children. Cattle are primary reservoirs for EHEC O157:H7 and the main source of infection for humans. Vaccination of cattle with different combinations of bacterial virulence factors has shown efficacy in decreasing EHEC O157:H7 shedding. It is, therefore, important to demonstrate whether vaccination of pregnant cows with EHEC O157:H7 induces high titers of transferable antibodies to avoid early colonization of calves by the bacteria. In this study we evaluated the ability of EspA, EspB, the C-terminal fragment of 280 amino acids of γ-intimin (γ-intimin C280) and inactivated Shiga toxin (Stx) 2 proteins to induce specific antibodies in colostrum and their passive transference to colostrum-fed calves. Friesian pregnant cows immunized by the intramuscular route mounted significantly high serum and colostrum IgG responses against EspB and γ-intimin C280 that were efficiently transferred to their calves. Antibodies to EspB and γ-intimin C280 were detected in milk samples of vaccinated cows at d 40 postparturition. Significant Stx2-neutralizing titers were also observed in colostrum from Stx2-vaccinated cows and sera from colostrum-fed calves. The results presented showed that bovine colostrum with increased levels of antibodies against EHEC O157:H7 may be obtained by systemic immunization of pregnant cows, and that these specific antibodies are efficiently transferred to newborn calves by feeding colostrum. Hyperimmune colostrum and milk may be an alternative to protect calves from early colonization by EHEC O157:H7 and a possible key source of antibodies to block colonization and toxic activity of this bacterium.

A systemic vaccine based on Escherichia coli O157:H7 bacterial ghosts (BGs) reduces the excretion of E. coli O157:H7 in calves.

Cattle are the main reservoir of enterohemorrhagic Escherichia coli O157:H7, a bacterium that, in humans, causes hemorrhagic colitis and hemolytic uremic syndrome (HUS), a life-threatening disease, especially in children and older people. Therefore, the development of vaccines preventing colonization of cattle by E. coli O157:H7 could be a main tool for an HUS control program. In the present study, we evaluated bacterial ghosts (BGs) of E. coli O157:H7 as an experimental vaccine against this pathogen. BGs are empty envelopes of Gram-negative bacteria, which retain the morphological surface make-up of their living counterparts and are produced by controlled expression of the cloned protein E, which causes loss of all the cytoplasm content. In this work, E. coli O157:H7 BGs were used for subcutaneous immunization of calves. The vaccinated animals elicited significant levels of BG-specific IgG but not IgA antibodies in serum. Low levels of IgA and IgG antibodies against BGs were detected in saliva from vaccinated animals. Following oral challenge with E. coli O157:H7, a significant reduction in both the duration and total bacterial shedding was observed in vaccinated calves compared to the nonimmunized group. We demonstrated that systemic vaccination with E. coli O157 BGs provides protection in a bovine experimental model. Further research is needed to reach a higher mucosal immune response leading to an optimal vaccine.

Profile of Shiga toxin-producing Escherichia coli strains isolated from dogs and cats and genetic relationships with isolates from cattle, meat and humans.

Pets can be reservoirs of Shiga toxin-producing Escherichia coli (STEC) strains. The aim of this study was to examine nine strains belonging to several serotypes (O91:H21, O91:H16, O178:H19, O8:H19, O22:H8, O22:HNT, ONT:H8), previously recovered from cats or dogs. To this end, we assessed a set of additional virulence genes (stx(2) subtype, subAB, ehxA, eae and saa), cytotoxic activity, and genetic relationships with strains isolated from cattle, meat and humans using pulsed-field gel electrophoresis (PFGE). Most of the isolates carried the stx(2) and/or stx(2vh-b) sequences, while only the O91:H21 isolate presented the mucus-activatable stx(2d) variant, as confirmed by sequencing the genes of subunits A and B. All the strains showed cytotoxic activity in cultured cells. One of the two O178:H19, selected for its high level of cytotoxicity in Vero cells, showed the ability to cause functional alterations in the human colon mucosa in vitro. None of the strains possessed the subAB, eae or saa genes and only the strains belonging to serotype O8:H19 carried the ehxA gene. The isolates shared 90-100% similarity by PFGE to epidemiologically unrelated strains of the corresponding serotypes recovered from cattle, meat or humans. Our results demonstrate that dogs and cats may have a role in the infection of humans by STEC, probably serving as a vehicle for bovine strains in the cycle of human infection, and thus emphasize the health risks for owners and their families.

Preovulatory follicle dynamics and ovulatory efficiency in sows with thrifty genotype and leptin resistance due to leptin receptor gene polymorphisms (Iberian pig).

This study compares follicular function and ovulatory efficiency in 20 sows with obesity/leptin resistance genotype (Iberian pig) and 20 females of lean commercial crosses (Large White × Landrace; LW×L). Estrous cycle was synchronized with progestagens; ovulation was induced with eCG and hCG, in half of the females of each group, to determine its effect. In females of both breeds not treated with gonadotropins, the number of follicles larger than 4.9 mm and the estradiol secretion increased throughout the follicular phase (P<0.05); estradiol values were similar at estrus detection (22.5±1.2 vs. 26.5±0.6 pg/ml respectively, for Iberian and LW×L sows). Moreover, ovulation rate was higher in Iberian pigs (15.3±1.3 CLs) than in LW×L (10.2±1.3 CLs; P<0.05), with mean progesterone values being 18.1±0.7 ng/ml in Iberian and 16.8±0.6 ng/ml in LW×L pigs. Thus, the preovulatory follicular growth and the ovulatory efficiency seem not to be the main limiting factors for reproductive efficiency in Iberian swine. The gonadotropins induced a significant increase, when compared to untreated females (P<0.05), in the number of follicles larger than 4.9 mm growing throughout the follicular phase; however, estradiol values at estrus were lower (P<0.05) in both breeds (9.2±0.7 pg/ml in Iberian vs. 8.6±0.8 pg/ml in LW×L), when compared with the nontreated animals, which suggests defective follicular function after gonadotropin stimulation. There were also no differences between genotypes in ovulation rate (15.2±1.3 vs. 12.7±1.8) and progesterone secretion (21.2±0.8 ng/ml in Iberian and 20.9±0.7 ng/ml in LW×L sows) in the treated animals. In conclusion, the current findings indicate that preovulatory follicular growth and ovulatory efficiency are not main limiting factors for prolificacy in a pig model of leptin resistance and obesity.

Genotypic and phenotypic characterization of enterotoxigenic Escherichia coli strains isolated from Peruvian children.

Enterotoxigenic Escherichia coli (ETEC) is a major cause of childhood diarrhea. The present study sought to determine the prevalence and distribution of toxin types, colonization factors (CFs), and antimicrobial susceptibility of ETEC strains isolated from Peruvian children. We analyzed ETEC strains isolated from Peruvian children between 2 and 24 months of age in a passive surveillance study. Five E. coli colonies per patient were studied by multiplex real-time PCR to identify ETEC virulence factors. ETEC-associated toxins were confirmed using a GM1-based enzyme-linked immunosorbent assay. Confirmed strains were tested for CFs by dot blot assay using 21 monoclonal antibodies. We analyzed 1,129 samples from children with diarrhea and 744 control children and found ETEC in 5.3% and 4.3%, respectively. ETEC was more frequently isolated from children >12 months of age than from children <12 months of age (P < 0.001). Fifty-two percent of ETEC isolates from children with diarrhea and 72% of isolates from controls were heat-labile enterotoxin (LT) positive and heat-stable enterotoxin (ST) negative; 25% and 19%, respectively, were LT negative and ST positive; and 23% and 9%, respectively, were LT positive and ST positive. CFs were identified in 64% of diarrheal samples and 37% of control samples (P < 0.05). The most common CFs were CS6 (14% and 7%, respectively), CS12 (12% and 4%, respectively), and CS1 (9% and 4%, respectively). ST-producing ETEC strains caused more severe diarrhea than non-ST-producing ETEC strains. The strains were most frequently resistant to ampicillin (71%) and co-trimoxazole (61%). ETEC was thus found to be more prevalent in older infants. LT was the most common toxin type; 64% of strains had an identified CF. These data are relevant in estimating the burden of disease due to ETEC and the potential coverage of children in Peru by investigational vaccines.