Pulsed-field gel electrophoresis and antimicrobial sensitivity testing were used as tools to investigate the epidemiology of Streptococcus uberis mastitis in dairy cows. A total of 62 different strains were found among 138 isolates from the four herds investigated, and between 10 and 26 different strains were found in each herd. There was no strain common to all four herds. Identical strains of S. uberis were detected from different quarters of individual cows and from cows within the same herd, suggesting that transmission from quarter to quarter and cow to cow had occurred. Despite the great variation in S. uberis strains, persistent infection with the same strain within a lactation was observed in most cows. Predominant strains were present in two herds. Preliminary investigations could not clarify why these particular strains might predominate, but in one herd there was a significant difference between the prevalence of clinical mastitis in quarters infected with the predominant strain and that in quarters infected with other strains, suggesting the greater virulence of the predominant strain. The wide variety of S. uberis strains found is consistent with an environmental source of S. uberis. However, evidence of direct transmission, the persistence of infection, and the predominance of particular strains in some herds indicate that S. uberis infections are epidemiologically complex and that the relative importance of these factors in the occurrence of mastitis may differ between herds.

A mouse-adapted strain of human enterovirus 71 (HEV71) was selected by serial passage of a HEV71 clinical isolate (HEV71-26M) in Chinese hamster ovary (CHO) cells (CHO-26M) and in newborn BALB/c mice (MP-26M). Despite improved growth in CHO cells, CHO-26M did not show increased virulence in newborn BALB/c mice compared with HEV71-26M. By contrast, infection of newborn mice with MP-26M resulted in severe disease of high mortality. Skeletal muscle was the primary site of replication in mice for both viruses. However, MP-26M infection induced severe necrotizing myositis, whereas CHO-26M infection caused only mild inflammation. MP-26M was also isolated from whole blood, heart, liver, spleen and brain of infected mice. CHO-26M harboured a single mutation within the open reading frame (ORF), resulting in an amino acid substitution of K 149 AI in the VP2 capsid protein; two further ORF mutations that resulted in amino acid substitutions were identified in MP-26M, located within the VP1 capsid protein (G 145 AE) and the 2C protein (K 216 AR). Infectious cDNA clone-derived mutant virus populations containing the mutations identified in CHO-26M and MP-26M were generated in order to study the molecular basis of CHO cell and mouse adaptation. The VP2 (K 149 AI) change was responsible only for improved growth in CHO cells and did not lead to increased virulence in mice. Of the two amino acid substitutions identified in MP-26M, the VP1 (G 145 AE) mutation alone was sufficient to increase virulence in mice to the level observed in MP-26M-infected mice.

To improve diagnosis of mastitis in dairy cattle, a multiplex polymerase chain reaction (PCR) assay was developed for the simultaneous detection of the four major bacterial causes of bovine mastitis, Staphylococcus aureus, Streptococcus agalactiae, Streptococcus dysgalactiae, and Streptococcus uberis. The target sequence was the 16S to 23S rRNA spacer regions. The performance of the assay was examined with 117 milk samples collected from a subclinically infected herd, and the diagnostic specificities and sensitivities of the multiplex PCR were compared with conventional culture. PCR was significantly more sensitive than culture for detection of S. aureus and S. uberis, but there were no significant differences in sensitivities between PCR and culture for the detection of S. agalactiae and S. dysgalactiae. The results suggest that this multiplex PCR assay could be used as an alternative method in routine diagnosis for rapid, sensitive, and specific simultaneous detection of S. aureus, S. agalactiae, S. dysgalactiae, and S. uberis in milk samples.

Eighty isolates of Riemerella anatipestifer representing 71 outbreaks of riemerellosis in Thailand between 1994 and 1999 were serotyped using the gel diffusion precipitin test. Based on the precipitation patterns, 25 serological profiles containing one to three antigenic determinants were recognized. Heat-stable antigens of the organism reacted with antisera raised against 16 known serotypes and an untypable strain 698/95. The most prevalent serotype appeared to be serotype 7, followed by serotypes 5, 10, 21 and 1. Further study demonstrated that the untypable strain probably represents a new serotype. Analysis of the polymerase chain reaction-amplified rrs genes for restriction fragment length polymorphisms verified the inclusion of strain 698/95 within the species R. anatipestifer and supported earlier work excluding strain 670/89, which had originally been designated the reference strain of serotype 20. Therefore, it is suggested that the strain 698/95 could be adopted as a replacement for the reference strain of serotype 20. Attention should be paid to strains with multiple antigenic factors as they may be useful for the preparation of vaccines.

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