Lucienne Garcia Pretto-Giordano

Streptococcus agalactiae is a major causative agent of streptococcosis in Nile tilapia (Oreochromis niloticus) and understanding its etiology is important to ensure the sustainable development of global tilapia farming. Our research group recently observed contrasting disease patterns in animals infected with two different S. agalactiae serotypes (Ib and III). To better understand the basis for these divergent responses, we analyzed the brain transcriptome of Nile tilapia following bacterial exposure. Our findings revealed significant variation in the expression of genes involved in immune (e.g., CD209 antigen, granulin, C-X-C motif chemokine 10, prostacyclin synthase, and interleukins) and neuroendocrine (e.g., mmp13a, mmp9, brain aromatase, and pmch) pathways. The serotype Ib strain seems promptly recognized by the host, triggering a potent inflammatory response, whereas the serotype III strain elicited a less immediate response, resulting in more pronounced central nervous system (CNS) symptoms and behavioral effects. To the best of our knowledge, this is the first study to show serotype-specific immune responses to S. agalactiae in Nile tilapia. These findings are important for advancing disease management and control strategies in aquaculture. Identifying different immune reactions triggered by serotypes Ib and III may assist the development of more specific approaches for preventive measures, early detection, and effective treatment against streptococcosis.

This study investigated the occurrence of selected pathogens of bovine respiratory disease in fetal pulmonary tissue of cattle and associated these with patterns of disease. Fetal pulmonary (n = 37) tissues were evaluated by histopathology; immunohistochemical assays identified intralesional antigens of bovine alphaherpesvirus 1 (BoAHV1), bovine viral diarrhea virus (BVDV), bovine parainfluenza virus 3 (BPIV-3), bovine respiratory syncytial virus (BRSV), and Mycoplasma bovis. Molecular assays were performed to amplify reproductive disease pathogens and bovine gammaherpesvirus 6 (BoGHV6) from 12 lungs. The 2 patterns of pulmonary diseases were interstitial pneumonia (12/37) and suppurative bronchopneumonia (1/37). The frequency of the intralesional antigens identified was BRSV (16.2%; 6/37), BVDV (13.5%; 5/37), BoAHV1 (8.1%; 3/37), M. bovis (5.4%; 2/37), and BPIV-3 (2.7%; 1/37). Interstitial pneumonia was associated with BRSV (n = 3), BoAHV1 (n = 3), and BVDV (n = 2); suppurative bronchopneumonia contained a Gram-positive bacterium and BVDV and BRSV. Reproductive pathogens detected included Leptospira spp., (n = 3), BVDV, Neospora caninum, and Brucella abortus (n = 2). BoGHV6 DNA was identified in the lungs of two fetuses with interstitial pneumonia. These findings suggest that these fetuses were infected transplacentally by several pathogens. The role of some of these pathogens herein identified must be further elucidated in the possible participation of fetal disease.

Streptococcus agalactiae, also known as Group B Streptococcus (GBS), is a Gram-positive bacterium that colonizes the gastrointestinal and genitourinary tract of humans. This bacterium has also been isolated from various animals, such as fish and cattle. Non-coding RNAs (ncRNAs) can act as regulators of gene expression in bacteria, such as Streptococcus pneumoniae and Streptococcus pyogenes. However, little is known about the genomic distribution of ncRNAs and RNA families in S. agalactiae. Comparative genome analysis of 27 S. agalactiae strains showed more than 5 thousand genomic regions identified and classified as Core, Exclusive, and Shared genome sequences. We identified 27 to 89 RNA families per genome distributed over these regions, from these, 25 were in Core regions while Shared and Exclusive regions showed variations amongst strains. We propose that the amount and type of ncRNA present in each genome can provide a pattern to contribute in the identification of the clonal types. The identification of RNA families provides an insight over ncRNAs, sRNAs and ribozymes function, that can be further explored as targets for antibiotic development or studied in gene regulation of cellular processes. RNA families could be considered as markers to determine infection capabilities of different strains. Lastly, pan-genome analysis of GBS including the full range of functional transcripts provides a broader approach in the understanding of this pathogen.

E. coli is the main bacteria isolated from infected uterus and bacterial endotoxin can lead to fatal endotoxic shock. Systemic inflammatory response syndrome (SIRS) precedes the endotoxic shock. Thus, early recognition of SIRS is important for patient treatment and prognostic. In Brazil, Gram positive bacteria are responsible for approximately 20% of all pyometra cases, and there is limited information about pathophysiology of shock and tissue injury. The aim of this study was to investigate the capacity of Gram positive bacteria to cause SIRS in bitches with pyometra. A prospective follow-up of 67 bitches with pyometra was performed, which were classified as SIRS + and SIRS- on admission. All bitches were surgically treated (ovariohysterectomy), uterine contents were collected in a sterile manner and the material was submitted to microbiological evaluation. Were identified in 55.2% of bitches E. coli (G1), 23.9% other Gram negative bacteria (G2) and 20.9% Gram positive bacteria (G3). The leukocyte profile, serum biochemistry and prevalence of SIRS were similar between the groups. It is concluded that Gram positive bacteria have the capacity to promote tissue damage and can lead the patient to death after SIRS and shock, as well as by E. coli and other Gram negative.

O grupo das tilápias é um dos mais importantes entre os peixes cultivados. O sistema de cultivo mais utilizado é o intensivo, caracterizado por alta taxa de estocagem de peixes tornando-os susceptíveis às doenças infecciosas. A estreptococose é uma das enfermidades que causam grandes prejuízos econômicos. O objetivo deste trabalho foi testar uma vacina de Streptococcus agalactiae inativado aplicada por banho de imersão (b.i) e desafio com cepa homóloga. Foram utilizados 421 tilapia do Nilo (Oreochromis niloticus) com peso médio de 38,38 gramas, distribuídas em dois tratamentos (T1 e T2) e um grupo controle. No T1, os peixes foram vacinados com uma dose da vacina por b.i., com concentração de 5,4 x 108 UFC mL-1. O T2 recebeu duas doses da mesma vacina por b.i., com intervalo de 25 dias. O grupo controle recebeu água ultrapura por b.i. Os peixes foram desafiados por via intraperitoneal (i.p.), com concentração de 3,0 x 108 UFC mL-1 43 dias após a primeira ou segunda dose da vacina, de acordo com o tratamento. Os peixes dos tratamentos e controle foram monitorados durante 16 dias após o desafio. Comparativamente ao grupo controle os resultados obtidos para o T1 foram: p = 0,0805, RR = 0,79 (IC 95%: 0,61 – 1,01) e RPS = 21%. Para o T2 foram: p = 0,0296, RR = 0,74 (IC 95%: 0,56 – 0,96) e RPS = 26%. A mortalidade após o desafio foi de 57 peixes (40,71%) no T1, 51 (38,06%) no T2, e 76 (51,7%) no grupo controle. Não houve diferença significativa entre T1 e T2 e p=0,7445. Este resultado permite concluir que a vacina testada por b.i. com uma e duas doses apresentou baixa eficácia na imunização das tilápias.

The pathogenesis of a Streptococcus agalactiae was evaluated in a three-period experiment. Two groups of 40 fishes were intraperitoneally (i.p.) challenged in each experimental period with different infective doses of the pathogen. Doses varied from 1.0 x 10(6) to 1.5 x 10(8) CFU/fish. One group of 40 tilapia i.p. injected with tryptic soy broth (TSB) was used as a control group in each period. Mortalities varied from 67.5% in group 8 (infective dose 1.0 x 10(6) CFU/fish) to 90.0% in group 1 (infective dose 1.5 x 10(8) CFU/fish). Significant differences in mortalities were found only between group 8 and each of the other groups, except group 5 (infective dose 6.0 x 10(6) CFU/fish; mortality 75.0%). The highest mortality coefficients were observed in days 1-2 after inoculation (accumulated mortality 44.4%), and a second peak of mortality occurred at days 6-7. Challenged fishes from all the groups showed alterations in behaviour and similar clinical signs. These were anorexia, lethargy, erratic swimming, exophthalmia and ascites. Macroscopically, skin hemorrhage, splenomegaly, hepatomegaly with organ paleness and visceral adherences were observed. S. agalactiae was re-isolated from all the fishes from the experimental groups submitted to bacteriological examination. The illness observed in tilapia naturally infected with S. agalactiae was experimentally reproduced in this study, and the clinical signs produced were similar to those reported from the natural infections.

Tilapia aquaculture is one of the fastest-growing segments of fish production in Brazil. Nile tilapia (Oreochromis niloticus) is largely cultivated in the state of Parana, where Streptococcus agalactiae is the cause of severe disease outbreaks. The objective of this paper was to evaluate an inactivated S. agalactiae vaccine in tilapia for the control of streptococcal disease outbreaks. Tilapia, weighing approximately 20 g each, were intraperitoneally (i.p.) inoculated with 0.1 mL of the vaccine at a dose of 2.0 × 10⁸ colony-forming unit (CFU) mL⁻¹. One group of tilapia (treatment 1) received one vaccine dose, and the other group of tilapia (treatment 2) received two doses, with an interval of 21 days. The control group was i.p. inoculated with 0.1 mL tryptic soy broth fish⁻¹. Immunized and control tilapia were i.p. challenged with 0.1 mL of 3.0 × 10⁷ CFU mL⁻¹ at 30 days post vaccination. The fish were monitored daily for disease signs and for mortality for 16 days post challenge. A statistically significant difference (P=0.0045) was found between the mortality of treatments 1 and 2. The value of relative per cent of survival of 83.6% and 96.4%, respectively, indicate that this vaccine was efficient in Nile tilapia.

Este trabalho teve como objetivo o estudo de fatores de risco associados à mastite bovina causada por Prototheca zopfii. Foram analisadas 13 propriedades leiteiras dos Estados do Paraná e de São Paulo, segundo os seguintes critérios de seleção: confirmação prévia de casos de mastite por Prototheca spp., triagem pela pesquisa de Prototheca spp. em tanques de expansão e latões e rebanhos com contagem de células somáticas acima de 5x105cel mL-1. As amostras coletadas consistiram de: leite, água, solo, fezes e swab de teteiras. Prototheca spp. foi isolada de amostras de leite dos quartos mamários com mastite clínica ou subclínica em uma propriedade e de amostras de leite e do ambiente em quatro propriedades, nas quais foi isolada em amostras de: água de bebedouro, abastecimento, esgoto, empoçada no piso de estábulo e sala de ordenha, solo de piquete e pasto, teteiras, fezes de bezerros e suínos. Do total de 383 vacas examinadas, Prototheca spp. foi isolada em 20 (5,2%) vacas, sendo caracterizada como P. zopfii em 18. Os fatores de risco associados à mastite causada por P. zopfii foram: criação das vacas a pasto, alimentação dos animais com pasto e silagem, realização de ordenha mecânica em estábulo, permanência das vacas após ordenha em piquete sem alimento, criação de suínos próxima às instalações dos bovinos, existência de cães, gatos e roedores, falta de higienização dos tetos com água, pré-imersão dos tetos em aplicador com retorno e sem a troca do anti-séptico, alimentação dos bezerros com leite de vacas com mastite clínica e serem as vacas da raça holandesa.

De 88 vacas primíparas, oriundas de quatro rebanhos leiteiros, foram colhidas 1985 amostras de leite, ao longo dos 120 dias pós-parto, das quais 457 (23,02%) apresentaram resultados microbiológicos positivos. Os estafilococos coagulase negativos (ECN) foram isolados em 316 (69,14%) amostras, Corynebacterium bovis em 56 (12,25%), estreptococos em 41 (8,97%) e estafilococos coagulase positivos (ECP) em 38 (8,31%). Mastite clínica foi detectada em nove (10,23%) vacas. No primeiro dia pós-parto, 57 (64,77%) animais e 114 (32,66%) quartos apresentaram exames bacteriológicos positivos. Até o décimo quarto dia, ocorreu um decréscimo acentuado no número de vacas e quartos infectados, que posteriormente tendeu a estabilizar. Os ECN foram as bactérias mais isoladas ao longo de todo experimento, enquanto o número de estreptococos decresceu acentuadamente nas duas primeiras semanas pós-parto. As infecções por C. bovis aumentaram progressivamente a partir do parto. Entre os ECN, predominaram o S. hyicus e o S. intermedius e, nos estreptococos, os do grupo C e D. A contagem média de células somáticas (CCS), nos quartos infectados, foi de 508,914/m, enquanto que, nos quartos negativos, foi de 73,942/m.