Lactococcus spp. has emerged as a pathogen that is affecting global aquaculture, with L. garvieae, L. petauri, and L. formosensis causing piscine lactococcosis. While antimicrobials are commonly used to treat diseases in aquaculture, reports of antimicrobial resistance in fish isolates are increasing. However, little is known about the susceptibility patterns of Lactococcus spp. strains isolated from native fish species in Brazil. This study aimed to assess the antimicrobial susceptibility of these strains and establish a provisional epidemiological cutoff value for L. garvieae using the normalized resistance interpretation approach. A total of 47 isolates were tested: 17 L. garvieae, 24 L. petauri, and 6 L. formosensis. The isolates were classified as wild-type (WT) or non-wild-type (NWT) based on inhibition zone diameters. Isolates classified as NWT for three or more antimicrobial classes were considered multidrug-resistant, and the multiple antibiotic resistance (MAR) index was calculated. The results revealed heterogeneity in antimicrobial resistance profiles, with higher resistance to trimethoprim/sulfamethoxazole and norfloxacin. Resistance to other antimicrobials, including florfenicol and oxytetracycline (approved for use in Brazil), varied according to the bacterial species. Lactococcus petauri (87.5%) and L. formosensis (66.7%) showed the highest multidrug resistance, compared to L. garvieae (11.7%), along with higher MAR index values. These findings suggest that multidrug-resistant strains could pose future challenges in the production of native species, underscoring the need for ongoing monitoring of antimicrobial resistance and responsible use of antimicrobials in aquaculture.

Lactococcosis has emerged as an economically and ecologically significant disease in aquatic animals worldwide. This study employed multilocus sequence typing (MLST) to investigate the genetic diversity of Lactococcus spp. strains from Brazilian fish species and evaluate their phylogenetic relationships with global isolates to elucidate potential epidemiological connections involving multiple host species and distinct geographic regions. A total of 55 isolates from different laboratories had their DNA extracted, followed by the amplification and sequencing of the internal fragments of seven housekeeping genes (als, atpA, tuf, gapC, gyrB, rpoC and galP). Sequence types (STs) and clonal complexes (CCs) were defined. An unrooted neighbor-joining phylogenetic tree was generated using allele profiles from this study and those previously reported from other aquatic animal species. The isolates comprised 29 STs (11 previously reported, 18 novel ones), which were grouped into species-specific CCs: CC5 (L. formosensis); CC4, CC17, CC62 (L. garvieae); CC24, CC29, CC97 (L. petauri). Considerable genetic divergence was observed, with L. formosensis and L. garvieae forming heterogeneous populations, while L. petauri was more homogeneous. These findings describe the MLST structure of the sampled isolates and should be interpreted as hypothesis-generating rather than population-level estimates of genotype prevalence. Phylogenetics confirmed groupings within the CCs and revealed additional phylogenetic clustering patterns. In conclusion, the Brazilian Lactococcus spp. strains analyzed in this study constitute a genetically diverse population based on their STs. MLST and phylogenetic analysis demonstrated genetic relatedness between the L. garvieae and L. formosensis isolates from this study and those from other aquatic animal species. In contrast, all the STs identified for L. petauri in this study were unrelated to the MLST lineages responsible for outbreaks in Brazilian Nile tilapia (Oreochromis niloticus) and North American rainbow trout (Oncorhynchus mykiss). This suggests that piscine L. petauri populations in the Americas evolved from distinct ancestral origins.

This study evaluated the susceptibility of three Amazonian fish species, Arapaima gigas, Brycon amazonicus and Colossoma macroporum to Lactococcus spp., through experimental infection, and performed a detailed examination of the pathological damage. While lactococcosis-causing bacteria have already been isolated from these fish species, their pathogenic role remains unconfirmed. Lactococcus formosensis, L. garvieae and L. petauri strains were used to intracoelomically infect juvenile Amazonian fish. Following infection, the fish were monitored for 15 days to evaluate clinical signs and mortality. Fish that died or survived until the end of the experiment underwent bacteriological and histopathological examinations. Clinical signs were observed in the L. garvieae-challenged A. gigas, with one fish dying at five days post-infection. The remaining experimental animals, regardless of fish species, did not die. Bacteriological examination confirmed re-isolation of asymptomatic animals in the L. garvieae-challenged A. gigas and L. formosensis-challenged C. macropomum groups. No bacterial growth or histological alterations were observed in the control groups nor in the groups infected with L. petauri. Microscopic examination revealed L. garvieae-induced fibrinoid-necrotic hepatitis, lymphohistiocytic myocarditis and myositis in A. gigas, while L. formosensis caused lymphohistiocytic pericarditis in C. macropomum. These findings demonstrate that A. gigas and C. macropomum are susceptible to L. garvieae and L. formosensis, respectively, and that they can become bacterial carriers. The distinct histopathological profiles—particularly myositis (L. garvieae) and pericarditis (L. formosensis)—highlight species-specific manifestations of piscine lactococcosis in Amazonian fish species.

Lactococcus spp. has emerged as a pathogen that is affecting global aquaculture, with L. garvieae, L. petauri, and L. formosensis causing piscine lactococcosis. While antimicrobials are commonly used to treat diseases in aquaculture, reports of antimicrobial resistance in fish isolates are increasing. However, little is known about the susceptibility patterns of Lactococcus spp. strains isolated from native fish species in Brazil. This study aimed to assess the antimicrobial susceptibility of these strains and establish a provisional epidemiological cut-off value for L. garvieae using the normalized resistance interpretation approach. A total of 47 isolates were tested: 17 L. garvieae, 24 L. petauri, and 6 L. formosensis. The isolates were classified as wild-type or non-wild-type (NWT) based on inhibition zone diameters. Isolates classified as NWT for three or more antimicrobial classes were considered multidrug-resistant, and the multiple antibiotic resistance (MAR) index was calculated. The results revealed heterogeneity in antimicrobial resistance profiles, with higher resistance to trimethoprim/sulfamethoxazole and norfloxacin. Resistance to other antimicrobials, including florfenicol and oxytetracycline (approved for use in Brazil), varied according to the bacterial species. Lactococcus petauri (87.5%) and L. formosensis (66.7%) showed the highest multidrug resistance, compared to L. garvieae (11.7%), along with higher MAR index values. These findings suggest that multidrug-resistant strains could pose future challenges in the production of native species, underscoring the need for ongoing monitoring of antimicrobial resistance and responsible use of antimicrobials in aquaculture.

Piscine lactococcosis is a disease caused by Lactococcus garvieae, L. petauri and L. formosensis. It is considered to be an emerging disease and poses a risk to global aquaculture production. Studies of the genetic diversity of Lactococcus spp. strains have epidemiological importance since they provide information on the genetic relationships between isolates and indicate geographic and temporal distribution. The aim of this study was to evaluate the genetic diversity of Brazilian Lactococcus spp. isolates obtained from native fish species using PCR-based DNA fingerprinting techniques. For this, 36 isolates were selected and submitted to gyrB sequencing, molecular serotyping, REP-, BOX-, and RAPD-PCR approaches. The discriminatory power and the congruence between the typing methods were calculated. After the correct taxonomic discrimination of the isolate, a total of 14, 5 and 17 isolates were reclassified as L. garvieae, L. formosensis and L. petauri, respectively. L. garvieae and L. petauri are related to serotype I, while L. formosensis strains were classified as serotype II. The typing methods tested are useful for evaluating the genetic diversity of isolates and a comprehensive indication of the diversity was found. Heterogeneous and homogeneous populations were observed for L. garvieae and L. petauri, respectively. RAPD-PCR demonstrated a greater discriminatory power for L. formosensis and L. petauri, while REP-PCR showed better results for L. garvieae strains, thus these methodologies are recommended for genotyping these bacterial species. In conclusion, Brazilian isolates of Lactococcus spp. obtained from native fish species constitute a genetically diverse population and the techniques used in this study allowed a better genetic characterization of the isolates.

Bacterial diseases cause significant losses in the aquaculture of Amazonian fishes (Arapaima gigas, Brycon amazonicus, and Colossoma macropomum). Lactococcosis-causing bacteria (LCB) have emerged as important pathogens in global aquaculture, and although they have already been isolated from these fish species, their pathogenic role remains unconfirmed. This study evaluated the susceptibility of three Amazonian fish species to Lactococcus spp. through experimental infection and performed a detailed examination of the associated pathological damage. Lactococcus formosensis, L. garvieae, and L. petauri strains were used to intracoelomically infect juvenile Amazonian fish. Following infection, the fish were monitored for 15 days to evaluate clinical signs and mortality. Clinical signs were observed in the L. garvieae-challenged A. gigas, with one fish dying at five days post-infection. The remaining experimental animals, regardless of fish species, survived the challenge. All mortalities and surviving fish at the end of the trial underwent bacteriological and histopathological examinations. Bacteriological examination confirmed bacterial re-isolation from asymptomatic animals in the L. garvieae-challenged A. gigas (2/7) and L. formosensis-challenged C. macropomum (3/7) groups using MALDI-TOF MS and multiplex qPCR analysis. No bacterial growth or histological alterations were observed in the control groups or in the groups infected with L. petauri. Microscopic examination revealed L. garvieae-induced fibrinoid-necrotic hepatitis, lymphohistiocytic myocarditis, and myositis in diseased or asymptomatic A. gigas, while L. formosensis caused lymphohistiocytic pericarditis in surviving C. macropomum. These findings provide preliminary evidence suggestive of susceptibility of A. gigas and C. macropomum to infection by L. garvieae and L. formosensis, respectively, highlighting their potential to act as asymptomatic carriers. Furthermore, the histopathological findings were suggestive of species-associated lesion patterns of piscine lactococcosis in Amazonian fish, rather than indicative of high mortality.