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Ozonized oils are antimicrobial agents obtained from the combination of ozone and unsaturated fatty acids of vegetables oils. The aim of the present study was to evaluate the antimicrobial effectiveness of ozonized olive oil (OOO) and ozonized venadillo oil (OVO) against Escherichia coli and Staphylococcus aureus. The antibacterial activity was conducted by the agar dilution method to determine the minimum inhibitory concentration (MIC) and the bacterial Log10 reduction. The lowest MIC (4.5 mg/mL) against E. coli was obtained when OOO and OVO were ozonized during 12 and 6 hours, with 2.5 Log10 of bacterial reduction, respectively; while, the lowest MIC against S. aureus (1.5 mg/mL) was obtained when OVO was ozonized during 6 hours, with 3.4 Log10 of bacterial reduction. The OOO reached peroxide values of 642.53 and 703.7 mmol-equiv/kg after 6 and 12 hours, respectively, while an 892.12 mmol-equiv/kg was obtained after 6 hours for OVO. Data reported here suggest that both ozonized oils are promising effective treatment for bacterial infections. [ABSTRACT FROM AUTHOR]

Sand fly salivary proteins are on the spotlight to become vaccine candidates against leishmaniasis and to markers of exposure to sand fly bites due to the host immune responses they elicit. Working with the whole salivary homogenate entails serious drawbacks such as the need for maintaining sand fly colonies and the laborious task of glands dissection. In order to overcome these difficulties, producing recombinant proteins of different vectors has become a major task. In this study, a cDNA library was constructed with the salivary glands of Phlebotomus perniciosus from Madrid, Spain, the most widespread vector of Leishmania infantum in the Mediterranean basin. Analysis of the cDNA sequences showed several polymorphisms among the previously described salivary transcripts. The apyrase SP01B and the D7-related protein SP04 were successfully cloned, expressed in Escherichia coli, and purified. Besides, recombinant proteins were recognized by sera of hamsters and mice previously immunized with saliva through the exposure to uninfected sand fly bites. These results suggest that these two recombinant proteins conserved their immunogenic properties after expression in a prokaryote system. Therefore, this work contributes to expand the knowledge of P. perniciosus saliva that would be eventually used for the development of tools for vector control programs. [ABSTRACT FROM AUTHOR]

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Abstract: From July 2009 to date, a leishmaniosis outbreak has occurred in the south-west of the Madrid region (Spain) and has already accounted for more than 450 human cases in an area that comprises a population of approximately 500,000. The causative agent is Leishmania infantum and the main vector in the area is Phlebotomus perniciosus. Although canine leishmaniosis prevalence in the focus is not higher than the average in the Madrid region, a wild reservoir – the hare – has been implicated. In this study, we examined the exposure of Leishmania reservoirs in the area: dogs, hares, and wild rabbits to sand fly bites using the detection of specific IgG antibodies against P. perniciosus salivary gland homogenate or recombinant salivary proteins. Hares collected in a green park adjacent to the focus (n =59) showed positive exposure to P. perniciosus bites in comparison to hares from a non-endemic area (Czech Republic, n =18). A significant positive correlation was found between IgG response to yellow protein rSP03B and salivary gland homogenate (r =0.902) and between apyrase rSP01B and salivary gland homogenate (r =0.710). Wild rabbits captured in the study area (n =21) presented higher anti-saliva antibody levels than negative control sera and their IgG response against recombinant salivary proteins were positively correlated with salivary gland homogenate (rSP03B: r =0.710; rSP01B: r =0.666). All sera of dogs from the focus (n =34) showed higher anti-saliva IgG levels than that of non-exposed dogs. Moreover, dogs protected against sand fly bites through the use of topical insecticides and sleeping indoors showed significantly lower antibody levels than the non-protected ones. Antibody response to all three recombinant salivary proteins tested showed positive correlation with salivary gland extract (rSP03B: r =0.858; rSP01: r =0.864; and rSP01B: r =0.861). Data confirmed the exposure of hares, rabbits and dogs to P. perniciosus bites in the context of an outbreak of human leishmaniosis in Spain, highlighting their involvement in Leishmania transmission by supporting their role as potential reservoirs. This novel methodology represents a promising tool for further epidemiological studies that would help to design better strategies for the control of leishmaniosis in this area and other foci. [Copyright &y& Elsevier]

Abstract: Sorption mechanisms of pineapple powders produced by vibro-fluidized drying (VFD), spray drying (SD), freeze drying (FD) or vacuum drying (VD) were interpreted through enthalpy/entropy variations with respect to equilibrium moisture content (EMC) at 20, 30, 40 and 50°C and water activity (a w ) between 0.059 and 0.907. Powders from SD and VFD were more hygroscopic and integral properties indicated higher stability at changing a w compared to powders from VD and FD. Powders showed an enthalpy driven zone with equal isokinetic temperature (T B1 =499.0±4.8K) due to their same composition, and an entropy controlled zone with different isokinetic temperature, T B2 (VD: 116.0±40.6K; FD: 118.3±24.1K; SD: 95.8±23.4K; VFD: 81.8±6.3K), caused by different microstructures resulting from different drying methods. Powders from SD and VFD presented larger entropic zones that shifted the minimum integral entropy to higher a w , improving storage stability. [Copyright &y& Elsevier]

A study was carried out to compare the infection rates of Leishmania donovani in Phlebotomus orientalis sandflies at different microhabitats of a VL endemic village in Gedarif state, Sudan. DNA extracts of 1078 P. orientalis sand fly females sampled by CDC light traps from indoor, outdoor, peri-domestic, and sylvatic sites, in three transmission seasons, March-June 2016-18, in Helat-Belo village, were subjected to independent PCR amplifications targeting Leishmania kDNA and the cpb gene followed by ITS1 region sequencing. Leishmania kDNA was detected in 1.4% of the 1078 P. orientalis females captured in the area. Two of these specimens showed a characteristic 741 bp band of L. donovani after cpb gene amplification. The DNA sequence of the ITS1 region of the parasites matched the ITS1 L. donovani genotype F. There were no signficant differences between rates of infection of L. donovani in P. orientalis captured at different sites. Blood meals found in infected flies origninated from human (5 specimens), cattle (4 specimens) and donkey (2 specimens). The finding of fresh cow and donkey blood in the infected flies suggests the possible role of these animals in the zoopotentiation and/or zooprophylaxis against VL. The study provides important information for VL transmission models and control programs in East Africa.

In 2009, a large outbreak of leishmaniasis, associated with environmental changes, was declared near Madrid (Spain), in which Phlebotomus perniciosus was the vector, whereas the main reservoirs were hares and rabbits. Analysis of isolates from humans, vectors and leporids from the focus identified the Leishmania infantum ITS-Lombardi genotype. However, multilocus enzyme electrophoresis (MLEE), the reference technique for Leishmania typing, and sequencing of the hsp70 gene, a commonly used marker, were not performed. In the present study, 19 isolates from P. perniciosus ( n = 11), hares ( n = 5) and rabbits ( n = 3) from the outbreak area, all characterized as ITS-Lombardi in previous studies, were analysed by MLEE and hsp70 sequencing. The hsp70 results confirmed that all the analysed strains are L. infantum . However, by MLEE, 4 different zymodemes of L. infantum were identified based on variable mobilities of the NP1 enzyme: MON-34 (NP1 100 , n = 11), MON-80 (NP1 130 , n = 6), MON-24 (NP1 140 , n = 1) and MON-331 (NP1 150 , n = 1). The relative frequency of these zymodemes does not correspond to their usual occurrence in Spain. Moreover, MON-34 and MON-80 were found in P. perniciosus , hares and rabbits for the first time. These findings continue to provide insights into the outbreak and call for further studies with a higher number of strains.

Visceral leishmaniasis (VL, kala azar), caused by Leishmania donovani, transmitted by Phlebotomus orientalis, is a serious systemic disease that causes high morbidity and mortality rates in Sudan and other parts of East Africa and the world. Despite progress in understanding the epidemiology of the disease in East Africa, little is known about the host preference of P. orientalis in kala azar endemic villages of Sudan, which have some of the highest VL incidence rates in the world. The present study used host choice experiments and blood-meal identification approaches to determine the host preference of P. orientalis in kala azar endemic villages in Gedarif state, eastern Sudan. In the host choice experiment, tent traps were used to compare the attractiveness of cows, donkeys, sheep and goats for host-seeking P. orientalis. In the blood-meal identification study, blood-fed P. orientalis females, captured inside houses and peri-domestic habitats, were subjected to molecular typing using cytochrome b gene (cyt b) amplification and sequence analysis. Cows and donkeys were the most attractive to blood-seeking P. orientalis, followed by goats. Similarly, the blood-meal analysis of P. orientalis showed that the vector preferentially feeds on cows, followed by donkeys, humans and goats. The human blood index of P. orientalis was 19.4% (42/216), indicating a high zoophilic habit of the vector, both inside and outside the houses. Although the order of host preference varied by location, it was clear that cows are the most preferred host of P. orientalis in the area. Results are discussed in relation to the role of domestic/livestock animals in VL zoopotentiation and zooprophylaxis. Inference is made on the potential impact of insecticide treatment of cows in control of the vector and the transmission of VL in Sudan and other parts of East Africa., (© 2023 Royal Entomological Society.)

Leishmania infantum is a protozoan causing cutaneous and visceral leishmaniasis in several regions of the world, including the Mediterranean basin. Phlebotomus perniciosus is one of the most important vectors of leishmaniasis in the countries of the western Mediterranean basin. Sand fly vector control by insecticides remains a useful tool in the framework of leishmaniasis control programs. Pyrethroids are the most widely used class of insecticides for sand fly control. There is currently a lack of information on the insecticide susceptibility and discriminating concentrations (DCs) of P. perniciosus. The aim of this study was to determine lethal concentrations (LC 50 , LC 95 , and LC 99 ) and DCs of deltamethrin and permethrin against two strains of P. perniciosus from Madrid region (Spain). According to WHO tube bioassay protocol 24-h mortality obtained after 1-h exposure to deltamethrin (0.0003%, 0.001%, 0.003%, 0.01%, 0.03%, and 0.1%) and permethrin (0.003%, 0.01%, 0.03%, 0.1%, 0.3%, and 1%) was recorded. The LC 50 , LC 95 , and LC 99 as well as their respective 95% confidence intervals values were calculated from the baseline data using maximum probability estimates of parameters and binary logistic regression analysis (QCal software). The 100% mortality was recorded from 0.01% of deltamethrin for both P. perniciosus strains and from 0.1% and 0.3% permethrin for Fuenlabrada and Boadilla strains, respectively. Final DCs of deltamethrin and permethrin of each P. perniciosus strain were determined based on setting this parameter at twice the minimum concentration of insecticide that kills 99% (LC 99 ) at the following percentages: Fuenlabrada strain (0.0582% deltamethrin and 0.2648% permethrin) and Boadilla strain (0.0406% deltamethrin and 0.2446% permethrin). The results indicate that both P. perniciosus strains are susceptible to deltamethrin and permethrin and can be used in susceptibility tests, although Boadilla strain offers more consistent results. Due to the scarce existing literature on insecticide DCs for sand flies and the different current procedures to determine their susceptibility to insecticides it is a priority to multiply efforts in order to develop standards for monitoring insecticide resistance in sand flies., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Human cases of Crimean-Congo hemorrhagic fever (CCHF) were first detected in Spain in 2016. National human and animal health authorities organized a large, multidisciplinary study focusing on ticks as sentinels to determine the nationwide distribution of ticks with CCHF virus. Ticks were collected from animals and vegetation, samples pooled (12,584 ticks; 4,556 pools), and molecular methods used to look for the virus. We detected the virus in 135 pools from most of the regions studied, indicating that it is widespread in Spain. We found sequences of CCHF virus genotypes I, III, and IV in the tick species collected, most commonly in Hyalomma lusitanicum, suggesting this tick has a prominent role in the virus's natural cycle. The red deer (Cervus elaphus) was the host that most frequently yielded positive ticks. Our study highlights the need for larger studies in Spain to ascertain the complete risk to public health.

Experimental infections of Phlebotomus (L.) perniciosus from a colony established in Madrid (Spain) carried out with the Leishmania (L.) infantum zymodemes MON-1, MON-24, and MON-80 isolated in Tunisia are reported here. Laboratory-reared female sand flies were experimentally fed via membrane feeding device on a suspension of L. infantum promastigotes in defibrinated rabbit blood (10 7 /ml). Engorged females were dissected at progressive time points postfeeding to observe the intravectorial cycle of different L. infantum zymodemes. Development in the sand fly midgut of L. infantum parasites to the infective metacyclic promastigotes and monitoring the forward progression of parasites to finally reach the stomodeal valve (SV) of the sand fly were assessed. All tested L. infantum zymodemes developed properly in P. perniciosus. Experimental feeding with suspensions of promastigotes of all zymodemes led to very heavy late-stage infections. MON-24 and MON-80 zymodemes colonized the (SV) of P. perniciosus earlier than zymodeme MON-1, 2 and 4 days, respectively. Metacyclic promastigotes were observed in all experimental infections. The study shows for the first time that colonized P. perniciosus is able to acquire, retain, and develop in its midgut the zymodemes MON-24 and MON-80 isolated in Tunisia and highlights the putative role of this sand fly species in the transmission of such zymodemes to mammalian hosts in this country. The ability of experimentally infected sand fly species to transmit by bite such zymodemes needs to be assessed.