1. Investigación

Free-living amoebae (FLA) are ubiquitous and many isolates have been shown to be infected with amoeba-resisting bacteria, as the example of Acanthamoeba and Legionella interaction. Due to the high environmental prevalence of Acanthamoeba. in the Castilian Plateau (Spain), the aims of this work were to investigate the occurrence of Acanthamoeba and other FLA in water from several sampling points from four Drinking Water Treatment Plants (DWTP) and to investigate the presence of Legionella spp. and other amoeba-resisting bacteria in biofilms in raw and finished water, taking into account that no legislation exists for this protozoa control. Acanthamoeba was detected at different sampling points, and sand filters seemed to contribute to amoebic enrichment. After ozonation, a temporary decrease in viable amoebae was observed. The genotypes detected were T3, T4, and T5, revealing the first report of genotype T5 in waters from this region. Moreover, Balamuthia mandrillaris, Vermamoeba vermiformis and Paravahlkampfia sp. were detected. Regarding Legionella, PCR detection in raw and finished water was higher than by agar culture, but even higher after Acanthamoeba co-culture. Also, Legionella's presence was higher in raw water than in finished water. The decrease of free Legionella observed from raw (27.5%, by PCR) to finished water (3.4% by PCR) contrasted with the increase of Legionella-infected FLA from raw (30.7%) to finished water (52%). At biofilm, free Legionella was not detected, and the percentage of infected FLA was low (3.8%). Legionella species identified in these samples were L. drozanskii, L. donaldsonii and L. feeleii. Additionally, Acanthamoeba co-culture led to the isolation of Pseudomonas aeruginosa, P. stutzeri, P. fluorecens, Achromobacter xylosoxidans and Stenotrophomonas maltophilia. The highly disseminated presence of Acanthamoeba and the detection of amoeba-resisting bacteria inside amoebae highlight the importance of developing methods for controlling FLA in order to limit human pathogenic amoeba-resisting bacteria survival to the water purification processes.

The present study analyzes at the national level, the presence of circulating Legionella in the artificial aquatic systems of different facilities of all of them state-owned centers throughout Spain for 12 months. 1754 water samples from various state-owned centers were collected from January to December 2014. Samples were collected from the cooling towers and evaporative condensers (CTC), and water distribution networks such as domestic hot water (DHW), cold water for human consumption (CW), sprinkler irrigation systems (SIS), fire sprinkler systems (FSS), and water from decorative fountains (DF). All these facilities are considered, according to current regulations, as potential amplifying systems for bacteria and possible sources of infection by the generation of droplets and aerosols. The isolation and counting of Legionella in water samples was carried out using microbiological culture following the international normative UNE-EN-ISO 11,731:2007 (ISO 11,731:1998) and UNE-EN ISO 8199:2008 (ISO 8199:2005).The quantification of Legionella colonization, the annual distribution, and the geographical distribution of the Legionella isolates recovered in the water were analyzed. Besides, molecular techniques were used for the characterization of the Legionella non-pneumophila isolates. Legionella was recovered from 15.79% of the analyzed water samples. High colonization was more frequently detected in water samples from CTC, DHW, CW, and DF. Regarding the geographic distribution, positive samples of Legionella were obtained in 14 of the 18 Spanish locations analyzed. Legionella non-pneumophila was the most prevalent and was isolated from water samples from 13 different geographical locations (72%). Legionella anisa and Legionella jordanis were the most frequently non-pneumophila species isolated. Legionella donaldsonii was isolated for the first time in the water distribution networks in Spain. Legionella pneumophila sg 2-14 was detected in 13 locations and Legionella pneumophila sg 1 in 11 locations. Therefore, our study concludes that the presence of Legionella pneumophila and Legionella non-pneumophila species in these systems can be a potential threat to public health and should be examined thoroughly with complementary techniques, such as molecular techniques as a screen for routine diagnosis.

Background: HTLV-1 infection is a neglected disease, despite producing neurological and lymphoproliferative severe illnesses and affect over 10 million people worldwide. Roughly 5% of HTLV-1 carriers develop Adult T-cell leukemia/lymphoma (ATLL), one of the most aggressive hematological malignancies. Methods: A national HTLV-1 register exists since 1989 in Spain, a non-endemic country with a large migrant flow from Latin America and Equatorial Africa, where HTLV-1 is endemic. The main features of all patients diagnosed with ATLL in Spain up to date are reported. Results: A total of 451 cases of HTLV-1 infection had been reported in Spain until the end of year 2022. ATLL had been diagnosed in 35 (7.8%). The current average incidence of ATLL in Spain is of two cases per year. Women represent 57% of ATLL patients. Mean age at diagnosis was 47 years-old. Roughly 57% were Latin Americans and 26% Africans. At diagnosis, the majority presented with acute or lymphoma clinical forms. Survival was shorter than one year in most of them. Mean HTLV-1 proviral load was significantly greater in ATLL patients than in asymptomatic HTLV-1 carriers (2,305 vs 104 copies/104 PBMC). HTLV-1 subtyping in 6 ATLL patients found the 1a transcontinental variant (n = 4) and the Japanese variant (n = 2). All ATLL patients were negative for HIV-1, did not develop HTLV-1-associated myelopathy and were not transplant recipients. Conclusion: The rate of ATLL is very low in Spain and mostly associated to migrants from HTLV-1 endemic regions. Given the poor clinical outcome of ATLL, HTLV-1 testing should be performed at least once in all migrants coming from HTLV-1 endemic countries and in natives who have lived in or had sex partners from such regions.

Background: HTLV-1 infection is a neglected disease. Over 10 million people are infected worldwide, with hot spots of high endemicity across all continents. Roughly 5% of HTLV-1 carriers develop HTLV-1-associated myelopathy (HAM), a progressive subacute neurological disabling disease. Methods: We report the main features of patients diagnosed with HAM up to date in Spain, a non-endemic country with a relatively high migrant flow from Latin America and Equatorial Africa, where HTLV-1 is endemic. Results: A total of 451 cases of HTLV-1 had been recorded in Spain until the end of year 2022. HAM had been diagnosed in 58 (12.9%). The current incidence is of 2-3 new cases per year. Women represent 76%. Mean age at diagnosis is 49 years-old. Nearly 60% are Latin Americans. Although sexual transmission is the most likely route of HTLV-1 acquisition, up to 6 individuals had been infected following solid organ transplantation. Rapid onset myelopathy developed in all but one of these transplant recipients from three HTLV-1-positive donors. HTLV-1 subtype 1a transcontinental was the only variant recognized in HAM patients. HTLV-1 proviral load was significantly greater in HAM patients than in asymptomatic HTLV-1 carriers (677 vs 104 HTLV-1 DNA copies/104 PBMC; p = 0.012). Symptom relief medications and physiotherapy have been the only treatment providing some benefit to HAM patients. Neither significant clinical nor virological efficacy was noticed using antiretrovirals in at least 9 HAM patients. Two thirds of HAM patients ended up in a wheelchair and with urinary/fecal sphincter incontinence. Conclusion: HAM is the most frequent clinical manifestation of HTLV-1 infection in Spain, a non-endemic country. Middle aged women migrants from Latin America are the most frequently affected. Two thirds end up in a wheelchair despite using antiretroviral therapy.

Wild birds have been suggested to be reservoirs of antimicrobial resistant and/or pathogenic Enterococcus faecalis (Efs) strains, but the scarcity of studies and available sequences limit our understanding of the population structure of the species in these hosts. Here, we analysed the clonal and plasmid diversity of 97 Efs isolates from wild migratory birds. We found a high diversity, with most sequence types (STs) being firstly described here, while others were found in other hosts including some predominant in poultry. We found that pheromone-responsive plasmids predominate in wild bird Efs while 35% of the isolates entirely lack plasmids. Then, to better understand the ecology of the species, the whole genome of fivestrains with known STs (ST82, ST170, ST16 and ST55) were sequenced and compared with all the Efs genomes available in public databases. Using several methods to analyse core and accessory genomes (AccNET, PLACNET, hierBAPS and PANINI), we detected differences in the accessory genome of some lineages (e.g. ST82) demonstrating specific associations with birds. Conversely, the genomes of other Efs lineages exhibited divergence in core and accessory genomes, reflecting different adaptive trajectories in various hosts. This pangenome divergence, horizontal gene transfer events and occasional epidemic peaks could explain the population structure of the species.

More than a century ago, independent groups raised the possibility of using bacteria to selectively infect tumours. Such treatment induces an immune reaction that can cause tumour rejection and protect the patient against further recurrences. One of the first holistic approximations to use bacteria in cancer treatment was performed by William Coley, considered the father of immune-therapy, at the end of XIX century. Since then, many groups have used different bacteria to test their antitumour activity in animal models and patients. The basis for this reactivity implies that innate immune responses activated upon bacteria recognition, also react against the tumour. Different publications have addressed several aspects of oncolytic bacteria. In the present review, we will focus on revisiting the historical aspects using bacteria as oncolytic agents and how they led to the current clinical trials. In addition, we address the molecules present in oncolytic bacteria that induce specific toxic effects against the tumors as well as the activation of host immune responses in order to trigger antitumour immunity. Finally, we discuss future perspectives that could be considered in the different fields implicated in the implementation of this kind of therapy in order to improve the current use of bacteria as oncolytic agents.

The 2022 annual meeting of the HTLV & HIV‐2 Spanish Network was held in Madrid on December 14. We summarize here the main information presented and discussed at the workshop and review time trends for human retroviral infections in Spain. As transmissible agents, infections by human retroviruses are of obligatory declaration. Until the end of 2022, the Spanish national registry had recorded 451 cases of HTLV‐1, 821 of HTLV‐2, and 416 of HIV‐2. For HIV‐1, estimates are of 150 000 people currently living with HIV‐1 and 60 000 cumulative deaths due to AIDS. During year 2022, new diagnoses in Spain were of 22 for HTLV‐1, 6 for HTLV‐2, and 7 for HIV‐2. The last updated figures for HIV‐1 are from 2021 and counted 2786 new diagnoses. The slowdown in yearly infections for HIV‐1 in Spain points out that new strategies are needed to achieve the United Nations 95‐95‐95 targets by 2025. For the remaining neglected human retroviral infections, their control might be pushed throughout four interventions: (1) expanding testing; (2) improving education and interventions aimed to reduce risk behaviors; (3) facilitating access to antiretrovirals as treatment and prevention, including further development of long‐acting formulations; and (4) increasing vaccine research efforts. Spain is a 47 million population country in South Europe with strong migration flows from HTLV‐1 endemic regions in Latin America and Sub‐Saharan Africa. At this time universal HTLV screening has been implemented only in the transplantation setting, following the report of 5 cases of HTLV‐associated myelopathy shortly after transplantation of organs from HTLV‐1 positive donors. There are four target populations for expanding testing and unveiling asymptomatic carriers responsible for silent HTLV‐1 transmissions: (1) migrants; (2) individuals with sexually transmitted infections; (3) pregnant women; and (4) blood donors.