Periparturient blood T-lymphocyte PD-1 and CTLA-4 expression as potential predictors of new intramammary infections in dairy cows during early lactation (short communication).

BACKGROUND: The periparturient period in dairy cows is marked by immunosuppression which increases the likelihood of infectious disorders, particularly also mastitis. An in-depth understanding of peripartum leukocyte biology is vital for the implementation of highly successful post-partum disease prevention measures. Immune checkpoint molecules, such as programmed death 1 (PD-1) and cytotoxic T-lymphocyte antigen 4 (CTLA-4), are critical inhibitory receptors expressed on immune cells, particularly T cells, that drive immunosuppressive signaling pathways. However, the potential role of immune checkpoint molecules expression in T-cells on udder health has never been explored. Thus, the association between the occurrence of new postpartum intramammary infections (IMIs) and the expression of programmed cell death protein 1 (PD-1) and cytotoxic T-lymphocyte-associated antigen-4 (CTLA-4) on blood T-cells during the peripartum period was investigated. RESULTS: In this study, the incidence of IMIs by any pathogen in early lactation was not associated with a higher expression of PD-1 and CTLA-4 in the periparturient period. However, the incidence of IMIs by major pathogens throughout the first month of lactation was significantly associated with higher expression of PD-1 at 14 days before calving (P = 0.03) and CTLA-4 at parturition (P = 0.03) by blood T-cells. Also, the expression of CTLA-4 at D0 (P = 0.012) by T-cells was associated with the occurrence of persistent IMIs during the first month of lactation. CONCLUSIONS: To our knowledge, this is the first report to investigate the expression of PD-1 and CTLA-4 by blood T-lymphocytes during the periparturient period in dairy cows and to explore their relationship with the incidence of new IMIs in the postpartum period. Thus, a comprehensive understanding of leukocyte biology during peripartum would appear to be a prerequisite for the identification of resilient dairy cows or targets innovative (immunological) non-antibiotic approaches in the transition period.

Antimicrobial resistance and public and animal health risks associated with pathogenic Escherichia coli isolated from calves.

We aimed to determine the antimicrobial susceptibility profile of pathogenic Escherichia coli strains isolated from fecal samples of calves and buffalo calves (2008-2013), in Minas Gerais, Brazil, as well as the frequency of O157 gene and strains carrying extended-spectrum beta-lactamases (ESBL) and mobile colistin resistance (mcr) genes. E. coli strains (n=518) were tested for susceptibility against ten antimicrobials. Tetracycline was the antimicrobial with the highest resistance rate (382/518), followed by ampicillin (321/518), sulfamethoxazole/trimethoprim (312/518), chloramphenicol (192/518), gentamicin (126/518), ciprofloxacin (148/518), cefazolin (89/518), colistin (54/518) and cefoxitin (34/518). Multidrug resistance (MDR) was observed in 381/518 isolates. No strain harbored mcr or O157 genes, whereas 19/99 were ESBL positive. The most prevalent pathotype and phylogroup were STEC and B1, respectively. Age, EHEC pathotype and resistance to aminoglycoside and cephem were significantly associated with MDR in the multivariate model. Overall, E. coli strains showed high rates of resistance to penicillin, tetracyclines and folate inhibitors, in addition to an alarming rate of MDR and ESBL-producing strains.

Seroprevalence of Equine Leptospirosis in the State of Goiás, Brazil.

Leptospirosis, caused by Leptospira, is a zoonotic disease that, in horses, is linked to abortions, uveitis, and sporadic occurrences of liver and kidney disease, often resulting in significant economic losses for farmers. Research on the prevalence of leptospirosis in horses in the central-west region of Brazil has been relatively scarce. Thus, the present study aimed to determine the prevalence of leptospirosis in equine herds in the state of Goiás (Central Brazil). Blood samples were collected from 894 equids at 294 randomly selected farms divided into three different strata according to their herd characteristics. The microscopic agglutination test for the detection of anti-Leptospira agglutinins was carried out and the results showed that among the 294 sampled farms, 213 (72.9%; CI 95% 71.7-78.9) had one or more animals positive for leptospirosis, and of the 894 horses sampled, 513 (61.6%; CI 95% 54.3-69.0) were seropositive for leptospirosis. Djasiman, Icterohaemorrhagiae, and Australis were the most prevalent serogroups. The results showed a high prevalence of seropositive animals and a widespread distribution of positive farms in the state of Goiás. Thus, environmental sanitation measures and health education to prevent and control equine leptospirosis in the state are required.

Calpeptin is a potent cathepsin inhibitor and drug candidate for SARS-CoV-2 infections.

Several drug screening campaigns identified Calpeptin as a drug candidate against SARS-CoV-2. Initially reported to target the viral main protease (Mpro), its moderate activity in Mpro inhibition assays hints at a second target. Indeed, we show that Calpeptin is an extremely potent cysteine cathepsin inhibitor, a finding additionally supported by X-ray crystallography. Cell infection assays proved Calpeptin's efficacy against SARS-CoV-2. Treatment of SARS-CoV-2-infected Golden Syrian hamsters with sulfonated Calpeptin at a dose of 1 mg/kg body weight reduces the viral load in the trachea. Despite a higher risk of side effects, an intrinsic advantage in targeting host proteins is their mutational stability in contrast to highly mutable viral targets. Here we show that the inhibition of cathepsins, a protein family of the host organism, by calpeptin is a promising approach for the treatment of SARS-CoV-2 and potentially other viral infections.

Impact of an autophagy-inducing peptide on immunogenicity and protection efficacy of an adenovirus-vectored SARS-CoV-2 vaccine.

Because of continual generation of new variants of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), it is critical to design the next generation of vaccines to combat the threat posed by SARS-CoV-2 variants. We developed human adenovirus (HAd) vector-based vaccines (HAd-Spike/C5 and HAd-Spike) that express the whole Spike (S) protein of SARS-CoV-2 with or without autophagy-inducing peptide C5 (AIP-C5), respectively. Mice or golden Syrian hamsters immunized intranasally (i.n.) with HAd-Spike/C5 induced similar levels of S-specific humoral immune responses and significantly higher levels of S-specific cell-mediated immune (CMI) responses compared with HAd-Spike vaccinated groups. These results indicated that inclusion of AIP-C5 induced enhanced S-specific CMI responses and similar levels of virus-neutralizing titers against SARS-CoV-2 variants. To investigate the protection efficacy, golden Syrian hamsters immunized i.n. either with HAd-Spike/C5 or HAd-Spike were challenged with SARS-CoV-2. The lungs and nasal turbinates were collected 3, 5, 7, and 14 days post challenge. Significant reductions in morbidity, virus titers, and lung histopathological scores were observed in immunized groups compared with the mock- or empty vector-inoculated groups. Overall, slightly better protection was seen in the HAd-Spike/C5 group compared with the HAd-Spike group.

Bats (Mammalia: Chiroptera) in urban-rural interfaces: community structure associated with pathogen screening in São Paulo-the largest metropolitan region in Brazil.

Little is known about the influence of the urban environments on bat species 'ecology. The urbanization process potentially lead to critical ecological changes in bat communities' intra and interspecific pathogenic transmissions dynamics. To date, the monitoring of pathogens in bats in Brazil has only been done with bats found dead or alive in households, from rabies surveillance systems. The present work aimed to investigate how urbanization influenced bat richness, relative abundance and pathogen occurrence. Most captured bats were Phyllostomidae, especially Sturnira lilium, Artibeus lituratus, A. fimbriatus, Glossophaga soricina, and Platyrrhinus lineatus, among others. From preserved-rural towards urban areas the lesser the bat richness, the higher the relative abundance of the captured bats. Noise level, luminosity and relative humidity correlated with bat abundance. The proportion of genders, sexually active bats and their size (weight, right forearm length, and body condition index) were stable throughout the investigation. Still, the proportion of pregnant females was higher in Spring and the number of juveniles in Summer, evidencing the seasonality of reproduction. Several Enterobacteria were isolated, evidencing a significant role of bats in the circulation of pathogens of medical and veterinary interest. These results are crucial in the pursuit of a harmonious coexistence between humans, bats and domestic animals in areas with different levels of anthropization.

Economic analysis of bovine brucellosis control in the Rondônia state, Brazil.

Bovine brucellosis, mainly caused by Brucella abortus, is a worldwide distribution anthropozoonosis that causes great economic losses. In 2001, Brazil launched the National Program for the Control and Eradication of Brucellosis and Tuberculosis (PNCEBT). Contemporaneously, a great effort to characterize the epidemiology of the disease in Brazilian states was started. In the state of Rondônia, a first epidemiological study was carried out in 2004, revealing a prevalence of 35.2% of infected herds and 6.22% of seropositive females. In 2014, after a successful heifer vaccination program with strain 19 (S19), a second study detected a reduction in the prevalence of infected herds to 12.3% and of seropositive females to 1.9%. The present study aimed to quantify and compare the costs and benefits related to the control of bovine brucellosis in the state through an accounting analysis. Vaccinating heifers and performing serological tests to move animals were computed as private costs. The expenditures of the state official veterinary service for brucellosis control were considered public cost. The considered benefits of lowering prevalence were decreased cow replacement, decreased abortions, decreased perinatal and cow mortality, and increased milk production. Considering private and public costs, the net present value (NPV) was estimated at US$ 18.3 million, the internal rate of return (IRR) was calculated at 23%, and the benefit-cost ratio (BCR) was 1.7. When considering only the private costs, the NPV was US$34.9 million, the IRR was 49%, and the BCR was 3.0, meaning that the bovine producer had a return of 3 for each unit of currency invested. The results showed that the bovine brucellosis control measures implemented in the state of Rondônia, which had as its main strategy the vaccination of heifers with S19, produced highly advantageous economic results. The state should continue with its vaccination program, stimulating the use of the RB51 vaccine in addition to S19, to achieve further reductions in prevalence at low cost.

Equine Brucellosis: Current Understanding and Challenges.

Brucellosis in equines, including horses, donkeys, and mules, is characterized by abscesses in tendons, bursae, and joints. Reproductive disorders, which are common in other animals, are rare in both males and females. Joint breeding of horses, cattle, and pigs was found as the main risk factor for equine brucellosis, with the transmission from equines to cattle or among equines possible, although unlikely. Hence, evaluation of the disease in equines can be considered an indirect indicator of the effectiveness of brucellosis control measures employed for other domestic species. Generally, the disease in equines reflects disease status in the sympatric domestic species, mainly cattle. It is important to note that in equines, the disease has no validated diagnostic test, which limits the interpretation of available data. Finally, it is important to mention that equines also represent significant Brucella spp. infection sources for humans. Considering the zoonotic aspect of brucellosis, the significant losses due to infection, and the representativeness of horses, mules, and donkeys in the society, as well as the continuous efforts to control and eradicate the disease in livestock, in this review, we covered the various aspects of brucellosis in equines and compile the sparse and diffuse information on the subject.

Epidemiological Situation of Glanders in the State of Pará, Brazil.

Glanders is an anthropozoonosis caused by the bacteria Burkholderia mallei, affecting mainly equids. It has been eradicated in North America, Australia, and Western Europe, but continues to occur sporadically in countries in Asia, Africa, the Middle East, and South America. Its notification is mandatory by the World Organization for Animal Health. After 30 years, the disease reappeared in Brazil in 1999 and, thereafter, 1,413 outbreaks have been reported. However, the epidemiological situation of the disease in the country is not adequately known. Thus, 2718 animals from 654 properties in the state of Pará were randomly selected by sampling and examined using a serial protocol with Complement Fixation and Western Blot serological tests. The prevalence of properties infected with glanders in the state was estimated at 1.68% [0.84; 3.33] and of seropositive animals at 0.50% [0.27; 0.94]. The introduction of animals was individualized as a risk factor for disease introduction in the properties (OR = 5.9 [1.4; 25.5]). Despite the low prevalence of infected properties and seropositive animals, the state must review actions to fight the disease, considering that the strategies implemented have not affected the endemic balance of the disease. This process must involve all public and private agents interested in the topic.

A dietary spray-dried plasma feeding programme improves growth performance and reduces faecal bacterial shedding of nursery pigs challenged with enterotoxigenic Escherichia coli K88.

This study aimed to assess the effects of different spray-dried plasma (SDP) feeding programmes to pigs on performance, intestinal histomorphology and faecal bacterial shedding after an Escherichia coli K88 challenge. A total of 96 piglets (5.77 ± 0.01 kg) were weaned at 21 days of age (Day 0) and challenged with 3 ml of 1 × 1010 CFU of E. coli K88 in total 3.0 × 1010 CFU/animal on Days 0, 2 and 4. Pigs were fed nursery diets containing 0.0%, 3.0%, 6.0% or 9.0% SDP from weaning to 35 days of age; 0.0%, 1.5%, 3.0% or 4.5% SDP from 36 to 49 days; and the same control diet (without SDP), for the last 10 days of the experiment (50-59 days of age). Performance was measured from 35 to 59 days of age and faecal bacterial shedding and intestinal histomorphometry were evaluated at Days 28 and 49 of age respectively. From 21 to 35 days of age, there was a linear effect for body weight (BW) and average daily gain (ADG), a trend of linear effect for average daily feed intake (ADFI) and a quadratic effect for feed:gain ratio (FG). From 21 to 49 days, the 9.0:4.5% and 6.0:3.0% SDP feeding programmes improved BW, ADG and FG when compared to the other treatments. At 59 days of age, BW and ADG were increased by the two highest SDP feeding programmes. The 9.0:4.5% SDP feeding programme increased ADFI from 21 to 59 days of age, with 6.0:3.0% being intermediate and the other two treatments being lowest. The CFU counts of E. coli/g of faeces decreased linearly with increasing addition of SDP. These results indicate that an extended inclusion of increased SDP levels in post-weaning diets can improve growth potential and decrease bacterial shedding induced by E. coli K88.

Apparent prevalence and risk factors for bovine tuberculosis in the state of Paraná, Brazil: an assessment after 18 years since the beginning of the Brazilian program.

Bovine tuberculosis (bTB) impacts considerably animal production and one health worldwide. To describe the prevalence, risk factors, and spatial pattern of the disease in the state of Paraná, Brazil, a cross-sectional study was conducted from September 2018 to February 2019. The area was divided into seven regions. Within each region, farms were randomly selected, and a predetermined number of cows was selected and tested by a comparative cervical tuberculin test. 17,210 animals were tested across 1757 farms. Herd prevalence of bTB-infected herds in Paraná was 2.5% [1.87-3.00%]. It has varied from 0.8 to 3.98% among seven regions, with clustering being detected in the west, central, and northeast areas. Animal prevalence was 0.35% [0.21-0.59%] and has varied from 0.08 to 0.6% among the pre-set regions. No major shifts in the prevalence of bTB were detected since 2007. Large-sized herds, dairy production, and feeding with whey were detected to be correlated with the presence of bTB. Exclusively among dairy herds, veterinary assistance from cooperatives, possession of self-owned equipment to cool milk, and feeding with whey were correlated with the disease. Considering these results, it is recommended that the state of Paraná seek to implement a surveillance system for the detection of bTB-infected herds transforming them into free ones, if possible, incorporating elements of risk-based surveillance. Health education is also recommended to inform farmers about the risks of introducing animals without testing and of feeding raw whey to calves.

Production and Evaluation of Chicken Egg Yolk Immunoglobulin (IgY) against Human and Simian Rotaviruses.

Producing specific antibodies in chickens is an attractive approach for diagnosis or therapeutic applications. Besides the high immunoglobulin Y (IgY) yield transferred to the egg yolk and its suitability for large-scale production, such an approach is more bioethical for animal maintenance. The IgY technology offers new possibilities for application in human and veterinary diagnostics and therapeutics, including strategies for treating severe intestinal diseases in children, particularly in emerging countries. Herein, we describe the production and purification of polyclonal antibodies against rotavirus group A (RVA) in immunised hens aiming at its application in prophylaxis and treatment of rotavirus-induced diarrhoea. For this purpose, we inoculated Rhodia laying chickens (Gallus gallus domesticus) with two or three doses of RVA combined with adjuvants or only adjuvants (control group). As the egg-laying period began, the yolk protein purification processes yielded a high concentration of specific IgY, the highest titre resulting from the group of hens that received three doses of the immunogen. The purified IgY blocked the functional activity of RVA in MA-104 cells, thus confirming the neutralisation ability. Therefore, anti-RVA IgY could be a promising candidate for pre- and post-exposure prevention or treatment of rotavirus-induced diarrhoea.

Leptolysin, a Leptospira secreted metalloprotease of the pappalysin family with broad-spectrum activity.

Extracellular proteolytic enzymes are produced by a variety of pathogenic microorganisms, and contribute to host colonization by modulating virulence. Here, we present a first characterization of leptolysin, a Leptospira metalloprotease of the pappalysin family identified in a previous exoproteomic study. Comparative molecular analysis of leptolysin with two other pappalysins from prokaryotes, ulilysin and mirolysin, reveals similarities regarding calcium, zinc, and arginine -binding sites conservation within the catalytic domain, but also discloses peculiarities. Variations observed in the primary and tertiary structures may reflect differences in primary specificities. Purified recombinant leptolysin of L. interrogans was obtained as a ~50 kDa protein. The protease exhibited maximal activity at pH 8.0 and 37°C, and hydrolytic activity was observed in the presence of different salts with maximum efficiency in NaCl. Substrate specificity was assessed using a small number of FRET peptides, and showed a marked preference for arginine residues at the P1 position. L. interrogans leptolysin proteolytic activity on proteinaceous substrates such as proteoglycans and plasma fibronectin was also evaluated. All proteins tested were efficiently degraded over time, confirming the protease´s broad-spectrum activity in vitro. In addition, leptolysin induced morphological alterations on HK-2 cells, which may be partially attributed to extracellular matrix (ECM) degradation. Hemorrhagic foci were observed in the dorsal skin of mice intradermally injected with leptolysin, as a plausible consequence of ECM disarray and vascular endothelium glycocalyx damage. Assuming that leptospiral proteases play an important role in all stages of the infectious process, characterizing their functional properties, substrates and mechanisms of action is of great importance for therapeutic purposes.

Canine leptospirosis in stray and sheltered dogs: a systematic review.

The aim of this systematic review was to investigate the prevalence of leptospirosis among stray and sheltered dogs worldwide. Six databases were searched, which resulted in the retrieval of 476 articles. Sixty articles were selected for analysis according to 10 quality criteria. Among the selected papers, 26 papers [43.4% (26/60)] met five of the 10 quality criteria established, 10 papers [16.7% (10/60)] met three criteria, nine papers [15.0% (9/60)] met four criteria, six papers [10.0% (6/60)] met six criteria, four papers [6.7% (4/60)] met eight criteria, and three papers [5.0% (3/60)] met nine of the 10 criteria, whereas two papers [1.7% (1/60)] met two and seven [1.7% (1/60)] criteria. Publications originated mainly from the Americas [45.0% (27/60)] and in the last 16 years (2003-2019) [81.7% (49/60)], and most of the sampled dogs were stray dogs [65.0% (39/60)]. The most commonly used diagnostic test for leptospirosis was the microscopic agglutination test [78.4% (47/60)] followed by polymerase chain reaction [21.7% (13/60)], and the most common serovars were Canicola [71.4% (35/49)], Icterohaemorrhagiae [65.3% (32/49)], Grippotyphosa [40.8% (20/49)], and Pomona [40.8% (20/49)]. In conclusion, our results showed that Leptospira spp. are present in unowned dogs worldwide; however, the low-methodological quality of the recovered cross-sectional studies precluded a meta-analysis.

Leptospira sp. infection in bats: A systematic review and meta-analysis.

Bats are the only flying mammalian animals and are distributed worldwide. Bats are well-known hosts of several zoonotic viruses and bacteria, including Leptospira sp. Here, we performed a systematic review and a meta-analysis of evidence of Leptospira sp. infection in bats by examining studies published between 1964 and 2021. We reported the frequencies of various species and serogroups on all continents, several species and feeding habits of bats, and different diagnostic tools. Together, 33 papers from all continents with seven to 2077 individuals from one to 31 species were included. Molecular detection was conducted in most studies, followed by MAT (Microscopic Agglutination Test) and isolation and identification. Molecular characterization of Leptospira sp. revealed L. borgpetersenii as the most frequent species. Moreover, 179 positive samples for MAT contained the most likely infecting serogroups described, particularly the Australis serogroup. The percentage of positive tests in isolation and identification ranged between 0% and 0.5%. The highest frequency of Leptospira infection among the continents was observed in Asia, whereas South America had the lowest percentage. Finally, Nycteridae and Rhinonycteridae were the most frequently infected bat families. Our study provides valuable information about the epidemiology of Leptospira sp. infection in bats.

Improvement of the enrichment used in the EMJH medium (Ellinghausen-McCullough-Johnson-Harris) for the cultivation of Leptospira spp.

Changes were made to the original formulation of the EMJH medium (Ellinghausen-McCullough-Johnson-Harris) enrichment and some aspects such as growth time of Leptospira and utilization in the microscopic agglutination test (MAT) were evaluated and compared to the original enrichment and to a commercially available enrichment (DIFCO™). Leptospira samples (24 antigens) that make up our panel of antigens used in MAT were used, among them, reference and autochthonous strains isolated in Brazil. The samples were grown individually in the EMJH medium under the three previously mentioned conditions (adapted enrichment, original enrichment and commercial enrichment). In addition, 89 blood serums from domestic and wild animals were analyzed by MAT using the antigens grown in these media. All samples tested grew efficiently with the adapted enrichment, and the MAT results were satisfactory. Therefore, other laboratories could also benefit from the use of this adapted enrichment when culturing the Leptospira strains applied in their MAT panels.

Climate influence the human leptospirosis cases in Brazil, 2007-2019: a time series analysis.

BACKGROUND: Human leptospirosis is responsible for great losses and deaths, especially in developing countries, which can be mitigated by knowing the correct health indicators and climate influence on the disease. METHODS: Leptospirosis cases and deaths, population and precipitation were recovered from different databases (2007-2019). Annual incidence, mortality and case fatality rates (CFRs) of human leptospirosis and average precipitation were calculated for Brazil and its regions. Time series analysis using an moving average with external variable (ARMAX) model was used to analyse the monthly contribution and precipitation influence over leptospirosis cases for each Brazilian region and for the whole country. A forecast model to predict cases for 2020 was created for Brazil. RESULTS: Human leptospirosis exhibited heterogeneous distribution among Brazilian regions, with most cases occurring during the rainy season and precipitation influenced the disease occurrence in all regions but the South. The forecast model predicted 3276.99 cases for 2020 (mean absolute percentage error 14.680 and root mean square error 53.013). Considering the annual average for the period, the leptospirosis incidence was 1913 cases per 100 000 inhabitants, mortality was 0.168 deaths per 100 000 inhabitants and the CFR was 8.83%. CONCLUSIONS: The models built can be useful for planning leptospirosis surveillance and control actions for the whole country and its regions and, together with the health indicators, revealed no uniform epidemiological situation of leptospirosis in Brazil.

Seroprevalence and risk factors for bovine brucellosis in the state of Paraná, Brazil: an analysis after 18 years of ongoing control measures.

Seroprevalence and risk factors of bovine brucellosis (Brucella abortus) in herds and cattle were estimated by a cross-sectional study in the state of Paraná, Brazil. The state was divided into seven regions and a random, two-stage sampling was performed on properties and cattle from each region between 2018 and 2019. Serum samples were collected from 11,592 cows over 24 months from 1,757 properties and a questionnaire was applied to identify potential risk factors. As recommended by the National Program for the Control and Eradication of Animal Brucellosis and Tuberculosis (PNCEBT), serological testing for the detection of anti-Brucella antibodies included the buffered plate agglutination test (screening test) and the fluorescence polarization assay (confirmatory test). The seroprevalence of bovine brucellosis on properties and in cattle was 4.87% (95% confidence interval [CI]: 3.98-5.93%) and 2.24% (95% CI: 1.47-3.41%), respectively. Multiple logistic regression analysis identified larger herd size and failure to test for brucellosis as risk factors for the presence of anti-B. abortus antibodies. These results demonstrate no change in the prevalence when comparing initial studies conducted in 2002. Given our findings, it is recommended that policies for brucellosis control include a widespread vaccination program for higher prevalence areas and eradication approach to lower prevalence areas. All steps related to correct immunization of the herds should be verified and improved by training and education. Health education action must be carried out informing farmers about the risks of introducing animals not tested for brucellosis into their herds and the benefits of testing their herds regularly.

Virulence Genes Profile and Antimicrobial Susceptibility of Community-Acquired Bacterial Urinary Tract Infections in a Brazilian Hospital.

Urinary tract infections (UTI) are one of the most common diseases worldwide and Escherichia coli is the most common causative bacteria. Empirical treatment is challenging due to antimicrobial or multidrug-resistance. The aims of this study were to determine the uropathogens and their antimicrobial susceptibility profile, as well as to identify the phylogroups and virulence genes of E. coli strains, associated with community-acquired UTI in outpatients admitted at a Brazilian Hospital in southeast Brazil. In total, 47 bacterial strains were isolated from 47 patients, 44 women and 2 men (no gender record from one patient). The age of the patients whose urine culture were positive varied from 0 (less than one month) to 104 years. Most of the isolates were E. coli (41/47), followed by Klebsiella pneumoniae (2/47), Klebsiella variicola/Klebsiella aerogenes (1/47), Pseudomonas aeruginosa (1/47), Proteus mirabilis (1/47), and Citrobacter koseri (1/47). Most E. coli strains were classified as phylogroup B2 (15/41 = 36.59%) and B1 (12/41 = 29.27%) and the most common virulence genes among E. coli strains were fimH (31/41 = 75.61%), iutA (21/41 = 51.22%), and tratT (16/41 = 39.02%). Among the E. coli strains, 59% were multidrug-resistance and strains that were ampicillin, sulfamethoxazole/trimethoprim, or tetracycline-resistant exhibited more chance to be multidrug-resistance, with an odds ratio of 100.00 [95% confidence interval (CI) 9.44-1059.26], 22.50 (95% CI 3.95-128.30), and 12.83 (95% CI 2.68-61.45), respectively. Our results showed that E. coli was the main etiological agent identified and demonstrated high frequency of multidrug-resistance and virulence factors in bacterial strains isolated from UTIs.

Sperm viability, serological, molecular, and modified seminal plasma agglutination tests in the diagnosis of Leptospira in the semen and serum of bovine bulls.

The present study investigated the serum microscopic agglutination test (MAT) among 203 bovine bulls with reproduction by natural means, without apparent signs of orchitis or inflammation of accessory reproductive glands. Simultaneously, the semen of all bulls was subjected to sperm viability analysis and PCR based on the 16S rRNA gene. PCR-positive results of semen samples were confirmed by sequencing. A modified seminal plasma agglutination (MSPA) test, replacing the blood serum of all bulls in the MAT with seminal plasma was performed as well. Eight (8/203 = 3.9%) semen samples from bulls were considered nonviable (necrospermia and azoospermia) without relation to the PCR diagnosis. No agglutinin titers were identified in MSPA test. A high frequency (132/203 = 65%) of leptospiral agglutinin titers was identified in the MAT, particularly for the Sejroe serogroup (Hardjo CTG, 100/203 = 49.3%; Wolffi 74/203 = 36.4%; Guaricura 72/203 = 35.5%; and Hardjoprajitno 56/203 = 27.6%). Three (3/203 = 1.5%) semen samples of bulls were positive in the PCR, but these results were not confirmed by sequencing. The high frequency of serovars from the Sejroe serogroup typically adapted to bovines indicates the need for measures for the prophylaxis/control of the pathogen on the sampled farms. Discrepancies among the MAT, sperm viability, and molecular detection of leptospires in semen highlight the need for a combination of methods to diagnose leptospirosis in bovine bulls. To our knowledge, modified seminal plasma agglutination is described for the first time here to investigate anti-Leptospira antibodies produced locally in the genital tract in the diagnosis of bovine leptospirosis among bulls that reproduce by natural means.