Efficient production of acetoin by fermentation using the newly isolated mutant strain Lactococcus lactis subsp. lactis CML B4

With the aim of applying biotechnology to produce acetoin, a chemical that can be used as an aroma and as a building block for other compounds, several putative mutants with reduced lactic acid synthesis were obtained from a wild-type homolactic strain of L. lactis subjected to chemical mutagenesis. Among these mutants, a strain was isolated, CML B4, that showed reduced lactate dehydrogenase (LDH) and increased NADH oxidase (NOX) activities. Shaken flask cultures of this mutant strain mainly produced acetoin, increasing the levels produced compared to the wild-type strain by 15-fold. A point mutation detected in the ldh gene encoding LDH was probably the genetic defect responsible for this phenotype. In pH-controlled aerobic batch fermentation, the CML B4 strain produced more than 40 gL−1 acetoin, which was increased by up to 59 gL−1 in fed-batch fermentations, with yields close to 88 and 74%, respectively, and productivities exceeding 2 gL−1 h−1. These results indicate that this strain could be used industrially as a cell factory for the production of acetoin from bioresources.This work was supported by the Department of Industry, Trade and Tourism of the Basque Government [grant number S-PE05LE03]

Expression of a novel class of bacterial Ig-like proteins is required for IncHI plasmid conjugation

Antimicrobial resistance (AMR) is currently one of the most important challenges to the treatment of bacterial infections. A critical issue to combat AMR is to restrict its spread. In several instances, bacterial plasmids are involved in the global spread of AMR. Plasmids belonging to the incompatibility group (Inc)HI are widespread in Enterobacteriaceae and most of them express multiple antibiotic resistance determinants. They play a relevant role in the recent spread of colistin resistance. We present in this report novel findings regarding IncHI plasmid conjugation. Conjugative transfer in liquid medium of an IncHI plasmid requires expression of a plasmid-encoded, large-molecular-mass protein that contains an Ig-like domain. The protein, termed RSP, is encoded by a gene (ORF R0009) that maps in the Tra2 region of the IncHI1 R27 plasmid. The RSP protein is exported outside the cell by using the plasmid-encoded type IV secretion system that is also used for its transmission to new cells. Expression of the protein reduces cell motility and enables plasmid conjugation. Flagella are one of the cellular targets of the RSP protein. The RSP protein is required for a high rate of plasmid transfer in both flagellated and nonflagellated Salmonella cells. This effect suggests that RSP interacts with other cellular structures as well as with flagella. These unidentified interactions must facilitate mating pair formation and, hence, facilitate IncHI plasmid conjugation. Due to its location on the outer surfaces of the bacterial cell, targeting the RSP protein could be a means of controlling IncHI plasmid conjugation in natural environments or of combatting infections caused by AMR enterobacteria that harbor IncHI plasmids

Cardiorespiratory nematodes and co-infections with gastrointestinal parasites in new arrivals at dog and cat shelters in north-western Spain

Metastrongyloid and trichuroid cardiorespiratory nematodes of dogs and cats are considered emergent in several European countries, and an increase in the number and extent of endemic foci has been described. Since data on their prevalence are limited in this continent, faecal samples from new arrivals (365 dogs and 65 cats) at two animal shelters in North-western Spain were analysed using both floatation and Baermann techniques. In order to confirm the microscopic identification of Metastrongylidae first stage larvae, molecular characterization based on the sequence of the ITS-2 was performed. The possible influence of some variables such as the species, sex and age of the animals and the co-infection with other gastrointestinal parasites on the prevalence of cardiorespiratory nematodes was analysed. The most prevalent metastrongylid was Aelurostrongylus abstrusus (15.4%) followed by Angiostrongylus vasorum (4.1%) and Crenosoma vulpis (1.1%). Regarding trichuroids, Eucoleus aerophilus and/or Eucoleus boehmi eggs were detected in 28 dogs (7.7%) and four cats (6.2%). Almost all animals positive to cardiorespiratory nematodes (86.8%) were co-infected with gastrointestinal parasites. The prevalence of Metastrongylidae and respiratory trichuroids was significantly higher in dogs co-infected with Taenia spp. and Toxocara canis or Giardia duodenalis and Sarcocystis spp., respectively. In cats, a significant higher prevalence of Metastrongylidae nematodes was found in animals co-infected with Toxocara cati. Our results reveal that cardiorespiratory nematodes are common in companion animals from north-western Spain, showing higher prevalences than those previously reported from this country. This investigation represents the first report of C. vulpis, E. aerophilus and E. boehmi in dogs from Spain. The identification of a number of zoonotic parasites is of public health concern. Our results indicate that these nematodes should be included in the differential diagnosis of dogs and cats from north-western Spain showing respiratory or cardiac clinical signsThis research was funded by the Program for consolidating and structuring competitive research groups (ED431C 2019/04, Xunta de Galicia, Spain)S

The Age-Related Cryptosporidium Species Distribution in Asymptomatic Cattle from North-Western Spain

An age-related distribution of Cryptosporidium species has been reported in cattle, with C. parvum being predominant in suckling calves, C. bovis and C. ryanae being predominant in post-weaned calves and C. andersoni being predominant in adults. However, variants to this pattern have recently been reported. Thus, fecal samples (n = 594) from asymptomatic cattle were collected in north-western Spain. Animals were classified as 2 years (G5). Cryptosporidium detection and species identification were performed by SSU rRNA PCR. Individual Cryptosporidium prevalence was 16.7%; it significantly decreased with age. Cryptosporidium parvum was predominant in G1 and C. bovis was predominant in the rest of the age classes; C. bovis and C. ryanae were especially prevalent in G2 and G3. Cryptosporidium occultus was not found in suckling calves. Finally, C. andersoni and C. xiaoi were occasionally detected in G5. The presence of C. parvum in all age classes implies significant animal and public health concerns. The predominance of C. bovis in cattle older than 1 month supports the idea that the age-related pattern of Cryptosporidium species described in cattle is not fully consistent, and thus further studies are still needed to identify those factors determining the species distributionThis research was funded by Ministerio de Ciencia e Innovación (Spain), grant number AGL2016-76034-P and by Xunta de Galicia (Spain), grant number 2017-PG117S

Pedicle myiasis by Lucilia caesar (Diptera, Calliphoridae): an emerging disease in roe deer from north-western Spain

A total of 35 male roe deer (Capreolus capreolus) from Lugo province (north-western Spain) were examined for pedicle myiasis between 2020 and 2022. All these animals had died by different causes and were examined or preserved just after their death to avoid post-mortem infestations. After external and internal head inspection, five animals were diagnosed with severe myiasis at the basis of the antlers (14.29%; 95% confidence interval = 5.38–31.04). All the affected bucks presented a cutaneous wound (1.5–15 cm in diameter) around the pedicles with extensive tissue destruction, exposition of frontal and parietal bones, and massive infestation by dipteran larvae (2–12 mm long). Four of five roe deer showed whitish egg clusters adhered to the basis of the antlers. Moreover, in one animal, six larvae had penetrated the cranial cavity, and in two of them larvae were also found in the nasopharyngeal cavity. All the cases were recorded in summer (May–August). Morphological identification and subsequent molecular confirmation revealed that all animals were infested by different larval stages of Lucilia caesar (Diptera: Calliphoridae). This study represents the first report of pedicle myiasis in Spain. Since this disease is reported sporadically, the detection of five cases in a short period of time suggests an increase in the incidence of this myiasisS

Prevalence and distribution of infectious and parasitic agents in roe deer from Spain and their possible role as reservoirs

In order to provide up-to-date information about the prevalence of infectious and parasitic agents in Spanish roe deer (Capreolus capreolus), samples from 93 animals hunted from January 2013 to April 2015 were collected and analysed by parasitological, serological and molecular techniques. Sampled animals came from four roe deer populations corresponding to Oceanic, Continental, Mediterranean and Mountainous ecosystems of Spain. Data regarding sex, age and year were also considered. A high percentage of roe deer (95.7%) resulted positive for at least one agent. Sarcocystis spp. was the most frequently diagnosed genus (88.8%), followed by gastrointestinal nematodes (62.9%) and Schmallenberg virus (53.5%). Varestrongylus capreoli (38%), Anaplasma phagocitophylum (34.2%), Eimeria spp. (29.2%), Toxoplasma gondii (25%) and Cephenemyia stimulator (23.8%) displayed medium prevalences and, finally low percentages were registered for Moniezia spp. (6.7%), Dictyocaulus noerneri (2.4%) and Mycobacterium avium subsp. paratuberculosis (1.5%). No infections by Neospora caninum, Bovine herpesvirus, pestivirus or Coxiella burnetii were found. Climate was significantly associated with the prevalence of T. gondii, C. stimulator and A. phagocitophylum, with higher prevalences in animals from Oceanic and Mediterranean areas. Our results suggest that infections affecting Spanish roe deer, especially those of parasitic aetiology, represent one of the causes of the descent in the abundance of this ungulate in the last years. Moreover, the high prevalence of zoonotic agents such as T. gondii and A. phagocitophylum could also have a great relevance in the environmental and/or Public Health.This research was supported by a Programme for consolidating and structuring competitive research groups (GRC2015/003, Xunta de Galicia) and by the Research Project ‘RUMIGAL: Rede de estudo multidisciplinar dos ruminantes en Galicia’ (R2014/005, Xunta de Galicia, Spain)S

The Goat as a Risk Factor for Parasitic Infections in Ovine Flocks

Small ruminants in northwestern Spain are frequently managed in mixed flocks. Sheep–goat joint management has not been considered a risk factor for parasite infection, so the main objective of this study was to establish if mixed management with goats supposes a risk factor for parasitic infections in ovine flocks. Two thousand and ninety-three sheep were sampled from 74 commercial meat ovine flocks for diagnostic of the main parasites. Goat contact was a risk factor for sheep to be infected by protostrongylids, Dictyocaulus filaria, gastrointestinal nematodes and Eimeria spp. In contrast, Moniezia, Fasciola hepatica, Dicrocoelium dendriticum and Paramphistomidae prevalences were not influenced. Sheep–goat mixed management can be considered as a risk factor, since goats would act as a source of pasture contamination for interspecific parasites (protostrongylids, Dictyocaulus filaria and gastrointestinal nematodes). In relation to host-specific parasites, such as Eimeria spp., goats cannot be considered as a source for sheep, but competition for food and spaces between both small ungulates can suppose a reduced grazing area to sheep, provoking high environmental contamination and stress that facilitate their infection. Future epidemiological studies for parasitic infections in small ruminants should consider sheep–goat mixed management as a possible risk factor to be included in multivariate analysesThis work was supported by a program for consolidating and structuring competitive research groups (ED431D 2019/04, Xunta de Galicia, Spain)S

Impact of the COVID-19 Pandemic on the Evolution of Prevalence and Patterns of Cannabis Use among First-Year University Students in Spain—UniHcos Project

Among university students there has been evidence that the COVID-19 pandemic increased their psychological distress, exacerbated by social restrictions. The main objective of this study was to explore the impact of the COVID-19 pandemic on the prevalence and patterns of cannabis use among university students, in contrast to previous trends since 2012. Data from 10,522 first-year university students (73.3% female, Mage 19 (SD = 1.6)) from eleven Spanish universities collected between 2012 and May 2022 was analysed. Prevalences of cannabis use and their differences by sex were studied, as well as changes in patterns of use and its use for coping during the pandemic. It was found that during lockdown, all prevalence rates of cannabis use decreased in both sexes, showing no statistically significant differences and increasing again in the new normal period in both. Among regular cannabis users, 79.7% reported maintaining or increasing their cannabis use during the pandemic, and of these, half reported using cannabis to cope. Moreover, cannabis use in the usual household increased during the lockdown. These results show that although the overall prevalence of cannabis use was reduced during the lockdown, regular users tended to maintain or increase cannabis use. This could imply two different patterns of use among students, one social and occasional versus the other regular, providing new lines of research for prevention and the implementation of social policies.This research was funded by the National Drug Plan from the Ministry of Health, Social Services and Equality of Spain (Grant numbers: 2010|145, 2013|034 and 2020|030) and Instituto de Salud Carlos III (Grant number: PI16/01947)

Effectiveness of janus kinase inhibitors in relapsing giant cell arteritis in real-world clinical practice and review of the literature

Background A substantial proportion of patients with giant cell arteritis (GCA) relapse despite standard therapy with glucocorticoids, methotrexate and tocilizumab. The Janus kinase/signal transducer and activator of transcription (JAK/STAT) signalling pathway is involved in the pathogenesis of GCA and JAK inhibitors (JAKi) could be a therapeutic alternative. We evaluated the effectiveness of JAKi in relapsing GCA patients in a real-world setting and reviewed available literature.Methods Retrospective analysis of GCA patients treated with JAKi for relapsing disease at thirteen centers in Spain and one center in United States (01/2017-12/2022). Outcomes assessed included clinical remission, complete remission and safety. Clinical remission was defined as the absence of GCA signs and symptoms regardless of the erythrocyte sedimentation rate (ESR) and C-reactive protein (CRP) values. Complete remission was defined as the absence of GCA signs and symptoms along with normal ESR and CRP values. A systematic literature search for other JAKi-treated GCA cases was conducted.Results Thirty-five patients (86% females, mean age 72.3) with relapsing GCA received JAKi therapy (baricitinib, n = 15; tofacitinib, n = 10; upadacitinib, n = 10). Before JAKi therapy, 22 (63%) patients had received conventional synthetic immunosuppressants (e.g., methotrexate), and 30 (86%) biologics (e.g., tocilizumab). After a median (IQR) follow-up of 11 (6-15.5) months, 20 (57%) patients achieved and maintained clinical remission, 16 (46%) patients achieved and maintained complete remission, and 15 (43%) patients discontinued the initial JAKi due to relapse (n = 11 [31%]) or serious adverse events (n = 4 [11%]). A literature search identified another 36 JAKi-treated GCA cases with clinical improvement reported for the majority of them.Conclusions This real-world analysis and literature review suggest that JAKi could be effective in GCA, including in patients failing established glucocorticoid-sparing therapies such as tocilizumab and methotrexate. A phase III randomized controlled trial of upadacitinib is currently ongoing (ClinicalTrials.gov ID NCT03725202)