Physiological Races and Vegetative Compatibility Groups of Fusarium oxysporum f. sp. lactucae Isolated from Crisphead Lettuce in Japan

One hundred and sixteen isolates of Fusarium oxysporum f. sp. lactucae obtained from 85 fields in three crisphead lettuce-producing areas in Nagano Prefecture, Japan were typed for races using differential cultivars Patriot, Banchu Red Fire and Costa Rica No. 4. They were also grouped into vegetative compatibility groups (VCGs) using complementation tests with nitrate non-utilizing (nit) mutants. Two California strains reported as F. oxysporum f. sp. lactucum, a type culture of F. oxysporum f. sp. lactucae, and 28 avirulent isolates of F. oxysporum obtained from crisphead lettuce were included for comparison. Among Nagano isolates, 66 isolates were identified as race 1, and 50 as race 2. Race 1 strains derived from Shiojiri and Komoro cities and race 2 from Kawakami village and Komoro city. All isolates of race 2 were biotin auxotrophs, and the race could be distinguished based on its requirement for biotin on minimal nitrate agar medium (MM). Pathogenic isolates were classified into two VCGs and three heterokaryon self-incompatible isolates. Strong correlations were found between race and VCG. All the race 1 strains were assigned to VCG 1 except self-incompatible isolates, and all the race 2 strains to VCG 2. The 28 avirulent isolates of F. oxysporum were incompatible with VCG 1 and VCG 2. California strains was vegetatively compatible with VCG 1, and they were assigned to race 1. Based on vegetative compatibility, these two races of F. oxysporum f. sp. lactucae may be genetically distinct, and F. oxysporum f. sp. lactucae race 1 is identical to F. oxysporum f. sp. lactucum. Received 7 May 2002/ Accepted in revised form 6 September 2002     

G and P genotype profiles of rotavirus a field strains circulating in a vaccinated bovine farm as parameters for assessing biosecurity level

After improvement of hygiene protocols on boots in a bovine operation (farm A) in Ibaraki, Japan in September 2017, mortality of calves and the detection of 4 viral pathogen indicators, including bovine rotavirus A (RVA), became significantly low for one year. Subsequently, in the present study, these indicators and mortality were monitored and confirmed all were still low, except for the detection rate of bovine RVA in calves less than 3 weeks old. The present study aimed to investigate G and P genotypic profiles of RVAs in farm A from 2018 to 2020. Molecular analysis using semi-nested multiplex RT-PCR of positive RVAs (n=122) and sequencing of selected samples revealed the presence of G6, G8, G10, P[1], P[5] and P[11] genotypes and the prevalence of G and/or P combination and mixed infections. The most common combination of G and P types was G10P[11] (41.8%), followed by mixed infection with G6+G10P[5] (11.5%). Phylogenetic analysis of RVAs showed clustering with bovine and other animal-derived RVA strains, suggesting the possibility of multiple reassortant events with strains of bovine and others animal origins. Noteworthy as well is that vaccinated cattle might fail to provide their offspring with maternal immunity against RVA infections, due to insufficient colostrum feeding. Our findings further highlight the importance of RVA surveillance in bovine populations, which may be useful to improving effective routine vaccination and hygiene practices on bovine farms.     

Isolation and Molecular Characterization of Fowl Adenovirus and Avian Reovirus from Breeder Chickens in Japan in 2019–2021

Fowl adenoviruses (FAdVs) and avian reoviruses (ARVs) are ubiquitous in poultry farms and most of them are not pathogenic, yet often cause damage to chicks. A total of 104 chicken fecal samples were collected from 7 farms of breeder chickens (layers and broilers) in Japan from 2019 to 2021, and yielded 26 FAdV plus 14 ARV isolates. By sequencing, FAdV isolates were classified as FAdV-1, 5 and 8b. ARV isolates were classified as genotype II, IV and V. These results suggest that FAdVs and ARVs are resident in the breeder chicken farms in Japan.     

Synergistic effects of quaternary ammonium compounds and food additive grade calcium hydroxide on microbicidal activities at low temperatures

The microbicidal activities of mixtures of quaternary ammonium compounds (QACs) and food additive grade calcium hydroxide (FdCa(OH)₂) were evaluated in a suspension test at −20°C using an anti-freeze agent (AFA) containing methanol, or at 1°C, with varying contact time, toward avian influenza virus (AIV), Newcastle disease virus (NDV), fowl adenovirus (FAdV), avian reovirus (ARV), Salmonella Infantis (SI) and Escherichia coli (EC). At −20°C, the mixtures could inactivate AIV and NDV within 30 min, FAdV and ARV within 5 sec, and SI and EC within 3 min, respectively. AFA did not inactivate viruses and bacteria within 30 min and 10 min, respectively. At 1°C, the mixtures inactivated FAdV and ARV within 30 sec, AIV within 10 min, and NDV within 30 min. A mixture of slaked lime (SL) and QAC could inactivate FAdV and ARV within 30 sec, but could not inactivate AIV and NDV even after 60 min at 1°C. SL could not substitute FdCa(OH)₂ in order to exert the synergistic effects with QAC. Thus, QACs microbicidal activities were maintained or enhanced by adding FdCa(OH)₂. It is hence recommended to use QACs with FdCa(OH)₂, especially in the winter season.     

Evaluation of sprayed hypochlorous acid solutions for their virucidal activity against avian influenza virus through in vitro experiments

Hypochlorous acid (HOCl) solutions were evaluated for their virucidal ability against a low pathogenic avian influenza virus (AIV), H7N1. HOCl solutions containing 50, 100 and 200 ppm chlorine (pH 6) or their sprayed solutions (harvested in dishes placed at 1 or 30 cm distance between the spray nozzle and dish) were mixed with the virus with or without organic materials (5% fetal bovine serum: FBS). Under plain diluent conditions (without FBS), harvested solutions of HOCl after spraying could decrease the AIV titer by more than 1,000 times, to an undetectable level (< 2.5 log10TCID₅₀/ml) within 5 sec, with the exception of the 50 ppm solution harvested after spraying at the distance of 30 cm. Under the dirty conditions (in the presence of 5% FBS), they lost their virucidal activity. When HOCl solutions were sprayed directly on the virus on rayon sheets for 10 sec, the solutions of 100 and 200 ppm could inactivate AIV immediately after spraying, while 50 ppm solution required at least 3 min of contact time. In the indirect spray form, after 10 sec of spraying, the lids of the dishes were opened to expose the virus on rayon sheets to HOCl. In this form, the 200 ppm solution inactivated AIV within 10 min of contact, while 50 and 100 ppm could not inactivate it. These data suggest that HOCl can be used in spray form to inactivate AIV at the farm level.